Diversity and ubiquity of K channels

0306-4522/88$3.00 + 0.00 Pergamon Press pie 0 1988 IBRO

Neuroscience Vol. 25, No. 3, pp. 729-749, 1988 Printed in Great Britain

COMMENTARY

DIVERSITY

AND UBIQUITY

OF K CHANNELS

B. RUDY Department of Physiology and Biophysics, New York University Medical Center, New York, U.S.A. CONTENTS

1.

729

INTRODUCTION

2. CLASSES OF K CHANNELS

730

3. VOLTAGE-DEPENDENT K CHANNELS 3.1. Delayed rectifiers 3‘2. “A” currents

730 730 734

4. Ca-ACTIVATED K CURRENTS 5. INWARD RECTIFIERS

736 739

6. MODULATION OF K CHANNELS BY NEUROTRANSMITTERS AND SECOND MESSENGERS

739

7. SUMMARY AND PERSPECTIVES

74I

ACKNOWLEDGEMENTS

743

REFERENCES

743

1. INTRODUCFION

A K current was part of the original description of the action potential mechanism in squid giant axons by Hodgkin and Huxley.95 This current was shown to be responsible for the repolarization of the action potential. Later studies of the frog node of Ranvier revealed a similar mechanism for action potential generation, with a K current also responsible for repolarization. 67K currents have now been found in all cells which display action potentials mediated by Na, Ca, Na and Ca, or Cl. It has also become apparent that, in contrast to Na currents, K currents vary widely in their kinetics, voltage-dependence, pharmacology, single-channel behavior and other properties. Often various types of K currents are found in the same cell, demonstrating that their diversity is not strictly due to differences between cell types. Furthermore similar types of K currents can be found in different cells. The possibility of observing individual single-channels, and of separating each type of K current by kinetic, pharmacological, genetic and other experimental means, suggest that the diversity of K currents results from the presence of different types of K channels. As modem techniques such as the patch-clamp and channel incorporation in model membranes are applied to new cells, the list of K channels grows. Their diversity and prevalence is much larger than that observed for other ionAbbreviations:

AHP, afterhyperpolarization; 4-AP, Qaminopyridine; DTX, dendrotoxin; EGTA, ethylene~iycolbistamin~thylether) tetra-acetate; TEA, tetraethylammonium.

channels. Although K, Ca and even Na channels are found in non-excitable cells, K channels are the most ubiquitous and seem to be present in almost every eukaryotic cell. Interest in understanding K channels has grown over the last few years as awareness of their functional roles in excitable and non-excitable cells has expanded. In neurons, all K currents are basically inhibitory, as they provide the outward current which inward Na and Ca currents must overcome to produce depolarizing signals. As a result of this, K channels play an important role in regulating the level of neuronal excitability. The requirement for a large repertoire of firing patterns in nervous systems may therefore underlie the need for a diversity of K channels. Indeed, as targets of the actions of many drugs, toxins, neurotransmitters and second messengers, K channels have the capacity to underlie rich and ~diverse neuronal firing patterns and play a role in neuronal information coding and integration. This review stems from an interest in the molecular basis of’ K channel diversity and an attempt to classify tihe various types of K channels. As will be discussed a proper classification is not possible today. However, with t& molecular characterization of K channels and the advent of new specific toxins and drugs, a Iclassification may become possible. There iis a great deal of interest in neuroscience in specifying the characteristics that define a particular type of K channel. The possibility of recording intracelfular electrical activity from specific neurons in brain slices has greatly enhanced our understanding of the ionic basis of specific and varied neuronal

729

B. REEDY

730

responses (e.g. Ref. 122). K channels have often been found responsible for the electrical behavior of these neurons (e.g. Refs 99, 124, and other examples in 52). However, in most cases, it is not possible to carry out proper voltage-clamp studies and identify the currents directly. Rather, the ion-channel types responsible for a particular response are inferred from current injection studies. To make proper inferences from current-clamp data, it is imperative to understand the properties that define particular channel types. Various reviews of specific aspects of K channels have appeared ~~~~~~~y~2~8~25~57,80.92.113,115~137~165.'83.194 This review takes a broader view, focusing on properties that may allow distinction between channel types. It is shown that although there are some distinct K channel types, it is not always possible to assign every K current to a particular class. Thus, caution is stressed when identifying a channel type on the basis of a few, non-specific properties. Most of the examples used in the text, and particularly in the tables, are from voltage-clamp experiments where the properties of K currents were studied directly. The nomenclature used here is that used by most neuroscientists. While the basis for this nomenclature is purely historic, and it can be confusing, an improved terminology will require better understanding of channel types and the relationships between them. 2.

CLASSES OF K CHANNELS

Table I lists the properties of three general classes of K channeis: voltage-de~ndent channels (“delayed rectifiers”, and the “A” currents), the inward rectifiers, and the Ca-activated K channels. Subtypes of these families are discussed in further detail in the following sections. The table also illustrates properties of two neurotransmitter modulated K channels. This is further discussed in Section 6. 3. VOLTAGE-DEPENDENT 3.1.

K CHANNELS

Delayed rectifiers

The voltage-dependent K current in squid giant axons responsible for the repolarization of the action potential is activated with a delay upon membrane depolarization and rises slower than the Na current.94,95It is thus responsible for the delayed outward rectifying property of this membrane. Therefore, K channels responsible for currents like the one described by Hodgkin and Huxley are usually referred to as “delayed rectifiers”, These channels are widely distributed, being found in most nerve and muscle cells, as well as in many non-excitable cells. Two main criteria are used to consider some K channels as “delayed rectifiers”. The first is that the macroscopic current through these channels is similar in overall kinetic behavior and voltage-dependence to the K current described by Hodgkin and Huxley. The second criterion is one of exclusion. Voltage-

dependent potassium currents, activated by membrane depolarization, sufficiently distinct from typical “A” currents and which are not activated by a rise in the internal Ca concentration, are often called “delayed rectifiers”. However, “A” currents and Caactivated K currents can produce delayed rectification; and may also participate in action potential repolarization (see below). The result of the use of these criteria is that under this term are included voltage-dependent K currents with often very different inactivation properties, conductance-voltage curves, other kinetic properties, pharmacology and probably single-channel properties. At this point the term non-“A” voltage-dependent K channels may be a term that better describes the channels considered here. We will use the term “Z, ” to identify the current carried by this group of channels. Many cells, and most neurons, contain at least one type of “Ik”. Tabfe 2 compares the properties of selected “Z,” channels in various cells. As illustrated in the table, some cells may contain more than one type of “I,“. In the frog node of Ranvier, classically considered to exhibit a single K current, recent macroscopic kinetic and pharmacological studies have established that there are at least three types of K currents.” 57These findings show that an apparent homogeneous macroscopic current may contain more than one component. The three components of the current most likely represent distinct channel types because they are independent: it is possible to alter the properties of one without affecting the others. This would not be expected for a multi-state single channel type. At this point. the three components are considered “delayed rectifiers” because they all contribute in significant amounts to the total delayed rectifying current, all three are sufficiently distinct from typical “A” currents and none have been shown to be Ca-activated (see below). In frog and rat skeletal muscle, and in Aplysiu neurons, two components of “I,” have been described (see Table 2). Several components of non-“A” voltage-dependent K current are also observed in ~en~~~~ oocytes injected with total rat brain mRNA.YS” Other cells may also be expected to contain more than one type of “I,” channel. In squid giant axon, for example, K channel inactivation proceeds with two time constants and is incomplete even at high may be due to depolarizations. 4’ These properties either complex kinetics or the presence of several current components. as in the node of Ranvier. In squid axons, the complexity and incompleteness of (4-AP)‘i9.204 may block of “I K” by 4-aminopyridine also result, in part, from the presence of more than one current component. We must, therefore, view with caution the kinetic and pharamacological properties of the examples presented in Table 2. Not only was it assumed that “I,” was separated from other outward currents (such as Ca-activated K currents, etc.), but also that it was a discrete homogeneous current. At best these K currents may

in many cells.

Main K current

Functions: 1. Action potential repolarization and thus action potential duration. 2. Contributes to refractory period.

rties (in Single channel pro squid giant axon, $”et**lobster membranes% and skeletal muscle membranes:182 Single channel g ,-- 15-20 pS. Bursting behavior. Depolarization increases frequency of bursts.

Tl>K=_RB>NH,%Na.

Blocked by: Ca, Ba and internal TEA in most cells. 4-AP and external TEA in some cases. Ionic selectivity:

Widely distributed, with diverse kinetics, voltage-dependence and pharmacology.

& = g,n”k.

Inactivates slowly (hundreds of ms to s at room temp.), if at all, during a constant depolarization. Described by a first order kinetic equation raised to a power with or without a first order inactivating parameter:

Conductance increases upon membrane depolarization, rising sigmoidally after a depolarizing step and closing exponentially upon membrane repoladzation.

K channel

Voltage-dependent

Delayed rectifiers 8.14.25,92.94,95

Voltage-dependent

voltage-dependent

present in cells with rectifiers and other K

Functions: 1. Latency to first spike. 2. Regulation of firing rate. 3. Action potential repolarization?

Single channel properties: Single channel g = 15-20 pS. Does not show voltage-dependent bursting behavior like delayed rectifier. Opening probability decreases with time, with one or two time constants. 4-AP decreases mean open time and conductance.**lOs

(unknown cations).

KyNa for other monovalent

Ionic selectivity:

Blocked by 4-AP at mM concentrations. Blocked by Cs and TEA (in).

Present in several neuronal cells, muscle and eggs with more constant kinetics, voltage-dependence and pharmacology.

gA = g,a”k.

Conductance increases upon membrane depolarization. Inactivates fast during a step depolarization ( -=z100 ms at room temp). Operates around the resting potential: threshold for activation is close (usually negative to) the resting potential and steady-state inactivation is complete at the resting potential. Described by a similar equation as delayed rectifiers:

currents.

Usually delayed

Fast transient K currents.

K channels

as “Anomalous

by TEA (in),

Functions: 1. Provides resting g in some cells, dominating outward current at Ys close to rest. 2, Maintainanoe of long plateaux in action potentials. 3. Cardiac pacemaker activity. 4. Egg fertilization. 5. Regulation of firing frequency.

Single channel properties: In cardiac ventricular cells is - 20 pS. Single channel conductance is ohmic in both directions of rectification.“5.197

Tl>K>Rb>NH,%NNa.

Ionic selectivity:

Usually blocked Cs and Ba.

Present in skeletal and cardiac muscle, eggs of many animals and several vertebrate and invertebrate neurons.

Rectification is probably the result of block by intracellular Me ions and not a voltaeedependent gating; possib& by Mg competing with K for entry into the channel’s pore.“7

Conductance increases with hyperpolarization allowing K entry. Little outllow upon depolarization. Rectification depends on (K,,,) and it occurs around I’,.

Also known Rectifier”.

rectifiers

9.*0,81,84.92.130

I. K channels

Inward

Table Ca-activated

_

bv uM-

in cells. Functions: 1. Regulate Ca entry. 2, Action potential repolarization. Afterhyperpolarizations. Reguldt’lon~ of firing rate. Spike frequency adaptation, Burst termination.

Both widely distributed secretory and excitable

Tl:,K>Rb>NH,%Na.

Selectivity:

&ary~otoxin.2~.113.114.141.l66

II. Maxi-K,, or BK channel. Very large single channel conductance. Blocked by low concentrations of external TEA and by

I. SK channel. Small single channel conductance. Blocked by apamin.30.1”

Several types. Two main types:

In whole cell exneriments voltage and time dependence of the K currents includes the voltage and time dependence of Ca entry into the cell.

K channels activated or less internal Ca.

*9,0% .,.. Ic5?%ek3 . .I66

channe1”39,1W.‘79.~81

By closing upon serotonin action on the cell it mediates serotonin induced slow depolarization of resting potential and increase in duration of action potential which result in increase neurotransmitter release and presynaptic facilitation.

Present in Apfysia sensory neurons.

Turned-off by CAMP dependent protein kinase phosphorylation resulting from [CAMP] increases produced by serotonin.

Outwardly rectifying channel with weak voltage-dependence.

“S

Present in vertebrate central neurons and sympathetic cells.

Turned off by ACh acting on muscarinic receptors and several peptide neurotransmitters. Mediates excitation by these neurotransmitters.

Slow, “non-inactivating”, voltage-dependent K current, Activates close to rest or at hyperpolarized potentials.

Neurotransmitterand second messenger-regulated channels “M channel”4.5.7,8

4

G

f

1

3-20 (22°C)

l&20 (25°C) -50 (20°C)

> -40

> -40

_ -40

5. Bullfrog sympathetic neurons4.*

6. Rat sympathetic neuronsz2.48

7. Hippocampal neurons”’

8. Tritonia neurons’92.‘94

pyramidal

-30

1-s (20°C)

- -50

leg

4. Crab walking axons42.161

8840 (10°C)

IO-40 (6°C)

- -50

3. Myxicola giant axon27.74.‘b7

(88::C)

2fLlOO (12°C)

(12°C)

0.5-S

v -70

< -80

> -40

> -80

kinetics (ms)

2. Squid giant axon’3a.‘4.25.94.95

(c) I,, _ 18% in motor and sensory fibers

in motor and 52 in sensory 1

@I4~2 -27%

1. Frog sciatic node of Ranvierss5’ (a) Ikr, 55% of IK in motor and 30% in sensory fibers

Preparation

“threshold” (mV)

Activation

IL3 (10°C)

(220;)

(26 rn;,‘,SC)

(2%:)

-complete

“Slow”

r(l) = l-2 r(2)= 11 (10°C) 0.3 of I, no inactivatiot+’

>80

(12C:oZmV)

-4s (12’C. OmV)

(s)

Inactivation rate

Table 2. Delayed

(Ix)

10mM

ImM

block

N.B.

10mM

N.B. SmM

N.B.

N.B.

V-dep

V-dependent block

0.1 mM blocks 88% at OmV, 59% at 100mV204

N.B.

Kd - 1O-5 M

4-AP

rectifiers

10mM K,-8mM

Kd-

Block 3 mM

K,,-ImM

Block

Kd - 24 rnM’%

N.B. at 100mM’83

K,, > 0.3 mM

Kd - 0.3 mM

Blockers TEA out

Bl. 20pM capsaicin

Bl. 10nM Toxin Iz3

Others properties

1. Sigrnoidal X= 2

I. Sigmoidal 2. k,=O.S=

1. Sigmoidal

rise

rise -6OmV

rise

1. V-dep: 7-10mV e-fold incr. in g. 2. Sigmoidal rise in I. x = 223

1. Sigmoidal rise, n = 4 2. Present in class 1, 11, III axons

1. V-dep: 13mV e-fold change in g 2. Sigmoidal rise in I, x=2

1. V-dep: 2-S mV e-fold change in g. 2. Sigmoidal rise in I, n’,x =4. 3. k,, 5 = -50, -60 mV

1. Steepness of voltage dependence: fl rt2rs fl _ S-6 mV e-fold change in g 2. fl and t2 sigmoidal rise of I. 3. Is exponential rise 4. k, =-0.5 - -50, -6OmV fl + f2”4

Other

-50

I-10

>I00

Yes

>I00

N.B.

Bl. 4-16 mM?

Bl. 20mM

N.B. 30 mM

small effect 5mM -

“slow” seconds

Bl. 128mM >lOmM

Kd-

Block 126 mM

N.B. 10mM -

-

None in 10 s 5s, 1OmV

N.B. Cs or TEA,

I. Single channel g = 20 pS bursting kinetics 2. Bl. by Ba. 3. High selectivity K/Na. I. Sigmoidal rise

I. Sigmoidal rise.

1. Sigmoidal rise.

1. Sigmoidal rise 2. V-dep: 6 mV e-fold.

1. Slow I mainly in slow muscles

1. Fast: sigrnoidal rise, x=4

1. Sigmoidal rise 2. Similar in atria1 nodal, ventricular and Purkinje fibers

*I and activation rate incr. during development I. Sigmoidal rise 2. See Table 4 also

1. Sigmoidal rise

1. Sigmoidal rise

1. k, = 0.5 voltage at which there is 50% inactivation; 2. n factor at which activation parameter is raised to obtain signoidicity; 3. N.B. = not blocked; 4. N.I. = not inactivated.

20. Paramecium “o~‘45

19. Sacharomyces78

* -10

l&.50 (22°C)

-10

18. Coelenterate egg@

-20(-IOmV) (22°C)

* -40

17. Rat s~~atogeni~

(1) 6 (2) 60

N.B. BI. 44lmM Bl. >>40mM

Block N.B.

slow 0.2-0.3 IS-3

K,,m8mM

N.B. ?

Block 20mM

Block 70% 0.5 mM

Kd +. 5-10 mM

Bl. 40mM Bl. 40mM

>I-2s

Blocked 1mM75

N.B. 3 mM

N.B. IOmM

-

N.B.

-

l-2

S&150 20-80

cellist

-

?

N.I. in 10s

&,

WCS

-200ms

5-50

0.52 s (20°C)

> -40 > -25

N -50

> -70

u -50

2-10 or 0.8-2.5* ‘“,y.

30-50 (1SC)

(-40mV) 4-10 -60

16. Drosophila muscle adult, 4°C.“’ larva, 15”C,202

15. Rat skeletal muscle 2OoC58.59

Slow (20%)

Fas; (80%)

3”C.‘cl&U.uU

14. Frog skeletal muscle

13. Cardiac muscle, IK or I, ,?1*97JM

horizontal cells’”

12. Goldfish retinai

y -20

_ -40

11. Ambysroma spinal

culturesi

-20

-

10. Helix neurons’4*

IK(l) IK(2)

9. Aplysia bag cell~‘*~

734

B. RUDY

represent the properties of a majority component. However, their properties could be distorted if more than one component contributed significantly to the total current. Two K channel blockers, 4-AP and tetraethylammonium (TEA), are listed in Table 2. Both block 1‘ IK” channels to various degrees when applied externally. 4-AP is membrane permeable and blocks equally well applied from the inside or outside of a ce11’39.2a4although its blocking site is probably internal. TEA is impermeant and blocking effects differ when applied internally or externally, suggesting two types of binding sites.‘5.9’ Externally applied TEA blocks some but not other K currents.‘83 Internally applied TEA is less specific’*’ and usually less potent. It blocks many “Ik” channels, “A” currents and Ca-activated K currents, although at widely varying concentrations. Cs ions are another non-specific blocker of most K channels.92 Divalent cations, in particular Ba, also block “1,” channels, as well as other K currents.‘6.92.“3 These blockers, as well as Cs and internal TEA, will not be considered further. Table 2 illustrates the variations observed among some “Ik” channels. Although it is impossible to say at this point what determines the differences between these currents or how many “channel subtypes” they represent, we note the following patterns: 1. There is a K current with a very negative threshold, high sensitivity to 4-AP and relatively fast inactivation in the frog node of Ranvier, squid giant axon and skeletal muscle. In all these cases the membrane lacks a typical “A” current. Currents with similar characteristics have been observed in other cell types and labelled as an “A” current. Some of these are listed in Table 4. The distinction between this current and “A” currents is often difficult and is discussed in the next section; 2. The delayed rectifier in several types of vertebrate neurons, as well as in some molluscan cells, is characterized by a higher threshold than the currents just described. It is sensitive to external TEA and not to 4-AP. Many of these cells also show typical “A” currents; 3. Several cells show a very slow, exponentially rising, K current with a more negative threshold, e.g., in the node of Ranvier and Aplysia neurons (see Table 2) and also in skeletal muscle,” spinal motoneurons,‘*.19~‘72a and in Archidoris and Anisodoris neurons.‘58 We consider in a later section (see also Table 1) a voltage-dependent K current which is blocked by muscarine and called the “M” current. For purely historic reasons this current is usually not considered a “delayed rectifier”. Its properties, however, fall among the range of the properties of the currents considered here. Furthermore it will not be surprising if an “M” current is part of what is considered in some cases a “delayed rectifier”. We should note, however, that most of the currents described in Table 2 inactivate, albeit some do so very slowly, while “M” currents do not seem to inactivate.

3.2. “A” currents A relatively fast, transient K current which is activated by depolarizing steps from holding potentials negative to the resting potential was first described in molluscan neurons43,44.83*147 and termed “A” current, IA, by Connor and Stevens.43 IA in molluscan neurons has characteristics that distinguish this current from other K currents. IA activation is fast compared with other K currents and it also quickly inactivates. Steady-state inactivation is complete near resting potential (-40 mV) and the threshold for activation is lower than that of other K currents. Thus, this current operates in the subthreshold region for action potential generation, opening transiently with small depolarizations which start from hyperpolarized potentials. Such properties enable this current to regulate the frequency of repetitive firing when a neuron is spontaneously active or fires repetitively in response to tonic depolarization. The hyperpolarization that follows an action potential removes the steady-state inactivation of “A” current channels, which open transiently. The resultant transient outward current slows down the return of the membrane potential toward action potential threshold. As a result, the interspike interval is prolonged. The discovery of this current provided an explanation for the low frequency firing of certain crab leg axons which Hodgkin classified as Class I and II more than 20 years earlier.93 Indeed, Class I and Class II fibers were found to contain an “A” current which is lacking in Class III fibers.42 Another property of the “A” current, often used as a criterion for its presence, is its sensitivity to the convulsant 4-AP and its derivatives. In molluscan neurons, Thompson”’ showed that of three distinct K currents, a delayed rectifier, a calcium-activated current and an “A” current, the “A” current was most sensitive to these drugs. 4-AP blocked the “A” current with a Kd of approximately 2 mM. Currents with similar properties have been described in neurons of several species, including vertebrates, as well as in other cells. Table 3 compares the properties of “A” currents in various cells. In all these examples, many of the properties are similar to those that distinguish this current from other K currents in molluscan neurons: threshold for activation, rates of inactivation (calculated for temperatures around 2OC assuming a Qlo of 3) voltagedependence of steady-state inactivation, and sensitivity to 4-AP. However, as shown in the table, there is some variation among the various examples, particularly in voltage-dependence of inactivation and in inactivation rates. Using the latter parameter alone one may distinguish three groups of “A” currents in the examples of Table 3. In most vertebrate neurons the time constant of inactivation is around 50ms. This time constant is close to that obtained experimentally since many of those experiments were actually carried out at room temperature. I, in some molluscan and Drosophila neurons also fall in this

735

Diversity and ubiquity of K channels Table 3. “A” currents

Species Helix 14’ Anisodoris4’ Tritonia I92 Aplysia” Aplysia ‘03,‘88 Tritonia” Drosophila”‘”

Cell type

Activation “threshold” (mv)

Activation rate*t (ms)

Inactivation rate*tJ (ms)

-50 -60 to -50 -60 -50 -55 -60 -60

120 5-10


4-AI’ block -65 -75 -70

Crab’4z

visceral ganglion neurons ganglion neurons pleural ganglion neurons ink gland motoneurons bag cell neurons ventral swim intemeurons larval brain and ganglion neurons walking nerve Type I and II axons coxal receptor dendrite

Bullfrog”’

sacculus hair cells.

-50

10

50-80

95% at -65

Bullfrog4’ Rat2’.68 Rat’,,,‘,8 RaP Guinea-piglM

sympathetic neurons sympathetic neurons cultured hippocampal neurons cardiac ventricular cells dorsal root ganglion neurons

-60 -60 - 60 -40 -60

<5 very fast 3-10 2-10 4-20

50 55 10-40 20-40

RaP8

locus coeruleus neurons

-60

-

(1) 60-100 (2) 4.5 s -100

Rat@ Guinea-pig and rat” Ratus

cultured sensory neurons CA1 hippocampal cells

-50 -60

very fast

30-50 50

-75 -75 -70 -60 (1) -50 (2) -75 100% at -50 < -65 < -60

whole brain mRNA injected into Xenopus oocytes

-60

5-10

-50

-70

very fast 4-10 -10

40

-55 -70 to -75 -65 -75

80-100 ms -100

-60

0.5-l

-5

-60 to -65

-65

-1

-5

100% at -45

K,-2mM > 90% at 12 mM block block at 5mM block at 20 mM block at 5mM block at 20 mM K,>lmM K,,>lmM K,-2mM K,-2mM blocked at 5mM K,-1mM K,,-2mM

*Calculated for 20°C assuming a Q,, = 3. tRange of values is due in most cases to voltage-dependence. $Range of values is in some cases due to experimental variation. $Midpoint of steady state inactivation.

The time constant of inactivation in most molluscan neurons, however, seems to be somehow slower. The values given in the table may be in some cases an underestimate since they represent calculated values from experiments carried out at 4-5°C. Finally the “A” currents in crab axons and dendrite are much faster. “A” currents also differ on the voltagedependence of inactivation rates. For example Connor and Stevens found that the rate of inactivation of the current during a pulse was not voltagedependent. This, is also true for other examples in Table 3, but in others inactivation becomes faster with increased depolarization. Single-channel studies may reveal further diversity among the examples listed in Table 3. For example, the single-channel conductance of IA channels in rat cultured sensory neurons and guinea-pig dorsal root ganglions is around 20 pS (Table 1). A similar single-channel conductance is observed for “A” channels reconstituted in Xenopus oocytes injected with rat brain mRNA (Krafte and Rudy, unpublished observations). On the other hand the “A” channels in Drosophila brain neurons, although similar in macroscopic properties, have a single-channel conductance of about 5-8 pS.i8ia In current-clamp studies, sensitivity to 4-AP is often used as a criterion to identify a membrane property as an “A” current. Caution should be taken group.

in making such an interpretation, particularly in the absence of dose-response studies. As described in the previous section and below, other K currents are even more sensitive to 4-AP. Other currents, often considered “A” currents, that depart even more from the pattern of those examples in Table 3 are listed in Table 4. The first five examples differ from those in Table 3 mainly in that the inactivation and activation voltage-dependence is shifted towards more depolarized potentials. This may or may not affect the functional role of this current depending on the voltage-dependence of the inward currents. However, in Drosophila muscle, for example, in the Coelenterate egg and in retinal horizontal cells, the inward current is carried exclusively by Ca. The activation of these Ca currents occurs at more positive voltages. Therefore, the Z, in these cells may play a similar role as IA in the examples of Table 3. The shift in voltage-dependence could be an adaptation to the position of the threshold of the action potential on the voltage scale. These might be a “high-threshold” “ subclass” of “A” channels. The last examples in the table differ in two properties. The time constant of inactivation is much slower than that of the “A” currents in Table 3 and the current is much more sensitive to 4-AP. These two effects may be related to each other. Although 4-AP seems to block closed channels preferentially, since the

736

3.

RUDY

Table 4. Transient K currents which differ from typical “A” currents Species

Type

“Threshold” (mV)

1. Drosophila ‘12

pupal and adult muscle larval muscle

-35

2. Drosophila 202 3. Goidfish’90 4. Rabbit7a

retinal horizontal cells cardiac crista terminalis

5. Coelenterates5

egg cells

6. Rat’s5

sensory neurons nodose ganglion cardiac Purkinje fibres CA3 hippocampal neurons CA3 hippocampal Ventral photo-receptors

7. Sheep’“* 8. Guinea-pig76 9. Guinea-pig’” 10. Limulus”“~“9

on (ms) <5

-40

3-7

h 0.5* (mV) -45

-40

inact.? (ms)

4-AP block

*lO

blocked at IO mM

30-8

Y-dependent 500-1500 IO0

very fast

-30 -30

-25

<5

-41

-60

-

lMfo-2000

-40

-25

-60


-55

-

r,-500 or * 2500 t,*ms T:z+”s 600-1200 800

-25 -30

-70 -60 > -40

IO-80

-

blocked at IOmM blocked at 3 mM &--ImM blocked at 30 pM blocked at 0.5 mM blocked at _ 100 FM slow camp. blocked at 500 /1M blocked at 1mM

*Midpoint of steady-state inactivation. tApproximate time constant of current decline during a pulse at 20°C.

block is voltage-dependent and decreases with increasing depolarization,“9.2@’ Thompson”’ has suggested that there is also open channel block of “A” currents by 4-AP. We may propose that these currents with slower inactivation have a similar binding site for 4-AP, and since there is more time for open channel block, there is an apparent increase in affinity. Independent of what may be the relationship between this current and other “A” currents in Table 3, it is possible that we are dealing with two different and distinguishable current types, and that some cells contain both currents. This could explain, in part, the apparent discrepancies between various studies of transient outward currents. Hailiwell et ai.” describe a current in CA1 hippocampal pyramidal ceils that inactivates with a time constant of about 50 ms. Inactivation in cultured pyramidal cells is also fast.“’ Gustafsson et a1.76 in CA3 ceils describe a transient current which inactivates much more slowly. Examination of Figs 2 and 3 of Gustafsson et al. and Fig. 2 of Halliwell et al. suggests that in both cases there is a fast and a slow inactivating component. Furthermore, it appears in Fig. 3 of Gustafsson et al. that 100 pM 4-AP suppresses the slow component preferentially. The presence of a slow component, with somewhat different pharmacology, in addition to IA, was also noted by Mirolli’42 in a crab dendrite. Two transient components are also observed in Figs 3 and I I of Boyett’s’* study of sheep cardiac Purkinje fibers. The distinction between these two currents may help in clarifying the effects of dendrotoxin (DTX) and toxin I, two toxins derived from mamba snakes, which block transient K currents at very low concentrations. These toxins block the slowly inactivating transient K current in nodose ganglion sensory neurons’85 and IKp, in the frog node of Ranvier’” at nM or lower concentrations. Hailiwell et al.H7 found that higher concentrations of either toxin I or DTX were required to block the “A” current in CA1 hippocampal neurons. It is possible. that as is the case with 4-AP, “A” currents are less

sensitive to the snake toxins than the transient current which inactivates more slowly. Indeed Black and DoII~~ find two binding sites for DTX in chick brain synaptic membranes. The high-affinity binding site interacts strongly with B-bungarotoxin and is less abundant than the site with lower affinity. It is often not simple to distinguish between “A” currents and “delayed rectifiers”. For example, one of the components of the “delayed rectifier” in the frog node of Ranvier (Ixr,), the K current in squid giant axons, the prototype of “delayed rectifier” and the fast component of the “delayed rectifier” in frog skeletal muscle, seem very similar to the last examples shown in Table 4. The fast K current in rat skeletal muscle is actually almost as fast, and seems to have 4-AP sensitivity (see Table 2) similar to the examples of “A” current in Table 3 (see Section 7). Inactivation of K currents can be very complex and may depend on pulse pattern.13 One may have a single channel type with two inactivation rates or two channels. Caution should, therefore, be exercised when using inactivation as a unique criterion to define a channel type. 4. Ca-ACTIVATED K CURRENTS

The channels described above, “delayed rectifiers” and “A” currents, are voltage-dependent channels, that is, they are gated by the membrane potential. In a class of K-channels to be considered now, channel opening and closing depends on the Ca activity in the cell’s cytoplasm. In these channels an increase in the concentration of Ca inside the ceil leads to channel opening. Although in some cases there may be also voltage-dependent gating, for one of the Ca-activated K channel described below (the maxi-K Ca-activated channel or BK channel), the observed voltage-dependence of channel opening (e-fold increase in conductance per It&l 5 mV at ail Ca concentrations) is the result of voltage-de~ndent Ca binding to the channei.i43 However, we may leave open, at this point,

137

Diversity and ubiquity of K channels

least for the large Ca-activated channels, that Ca*+ is acting directly on the channel. Since their original discovery in molluscan neurCa-activated K currents have been found ons, ‘36~‘3* to be widely distributed, particularly in secretory and excitable cells (e.g. Refs 90, 113, 115, 137). Caactivated K currents play important functional roles. They are important in regulating firing frequency and if Ca accumulates upon repetitive firing, spike frequency adaptation. In some cells they also contribute to the resting potential. In secretory cells their activation results in the closing of voltage-dependent Ca channels, thus providing a negative feedback to regulate Ca entry. In this context they play an important role in the cell’s metabolism. This effect may also be important in neuronal and cardiac cells. In the case of secretory ceils there is evidence that the Ca activating the channel may either come from the outside (through Ca channels) or be released from internal stores.“*‘56,‘%Ca-activated K channels are frequent targets of modulation by neurotransmitters and second messengers (see Section 6). It is now clear that there are several types of

the question of whether some Ca-activated channels may also be gated directly by voltage. In addition, in whole cells, the Ca-activated K current shows a voltage-dependence derived from the gating of voltage-dependent Ca* + channels and the ensuing Ca* + entry. Ca* + affects many channels either through surfacecharge effects (see Ref. 92, Chapter 13) or through interactions with the gating machinery.‘* Ca-activation of channels, as considered in the examples given in this section, differs from these more general effects in that: (1) Ca*+ acts as a ligand to activate channel opening, much like some neurotransmitters activate their receptors; (2) Ca*+ acts only on the inside; (3) activation occurs at very low (micromolar or less) concentrations of internal Ca*+ (similar to those that can be expected physiologically). We should also differentiate a third mechanism by which Ca*+ can regulate K channels: that is by acting as a second messenger that can stimulate kinase C and Cacalmodulin kinases. These enzymes can modify K channel function (see Section 6). Reconstitution experiments in lipid bilayers clearly demonstrate, at

Table 5. Ca-activated K channels TEA-sensitive maxi-K(Ca)29,“~.‘3*.‘43,‘53 Slow AHP K(Ca130,96,‘55,‘” . I

Other names

BK channel,

Internal [Cal needed for activation

_ 10~6-10-5 M at voltages membrane potential

Voltage-dependence

Very voltage-dependent. G increases with membrane

maxi-K,,

I AHP, SK channel,

channel close to the resting

]OmS_lO-7 Ml0

Little or no voltage-dependence30,96.‘55

opening

Single channel conductance

15&250 pS (depending

on [K])

Pharmacology

Blocked Blocked

Tissue distribution (examples)

1. Chromaffin and secretion

increases

e-fold

lo-14 pS’O

by TEA,,,, Kd c 1 mM by nM charybdotoxinr4’ cells. 13*Regulates

TEA out resistant30.96.‘55.‘” Blocked by nM apamin30.37.96.107.155.166 Ca entry,

muscle.29~153~‘66Regulates excitability

Ca entry.

I. Bullfrog sympathetic neurons.‘55.‘9’ Slow AHP, spike frequency adaptation

2. Rat skeletal AHP

muscle.30,166 Long lasting

3. Bullfrog sympathetic cells.*~“~‘55 Spike repolarization. Fast AHP, firing frequency regulation

3. Hepatocytes. )’ Catecholamine increase in K flux

4. Anterior pituitary entry and secretion.

4. Neuroblastoma

cells.“’ Regulates Ca Oscillatory activity

6. Pancreatic

p-cells. 65~‘33~‘57 Regulates

7. Rat brain

membranes@ and tubular

Ca entry

membranes”4~‘43

induced

cells.96

5. Rat sympathetic neurons.” Regulation of firing frequency

5. Submaxillary and parotid glands.‘34 Regulates Ca entry, fluid secretion

8. Sarcolemma

Kc,

depolarization

Probability of channel for IO-15 mV29.‘43

2. Skeletal Modulates

apamin-sensitive

Slow AHP

738

B.

Ca-activated K currents. Two of these have been well characterized, and there is evidence that they are carried by two different types of channels. Table 5 summarizes the properties of these channels and gives examples of cells containing either. Both types are present in bullfrog sympathetic neurons”’ and skeletal muscle30’66 and they may co-exist in many other cells. Apamin, a component of bee venom,37 a component of Leiurus and charybdotoxin,‘4’ scorpion venom, each block one of the two types of Ca-activated K currents described in Table 5. This may provide very useful tools to identify these channels, particularly in current-clamp experiments where the properties of the channels cannot be obtained directly (however, see below). Some differences among BK channels of different cells have been reported. In muscle, the channel is blocked by internal TEA concentrations in the range of 30 mM or higher. 29,“4 In contrast the BK channel from rat brain&l or pituitary cells,‘@’ although otherwise similar, is blocked by much lower internal TEA. Furthermore, while the BK channel from brain may be modulated by kinases,20@“8’62 there is no evidence that this is the case in muscle. The macroscopic current through BK channels (1,) is typically a large, long-lasting outward current which is blocked by low concentrations of TEA and charybdotoxin. The current is very voltage-dependent even at constant Ca concentrations.29~34~‘55~‘66In con(Table 5) is a small current with trast to 1c, IAHp96~‘55,‘66 little voltage-dependence, higher Ca-sensitivity and is not blocked by TEA. The underlying single-channel (SK) has only been studied in muscle.” There is a diversity of “BK” channels. Model membranes where channels from rat brain synaptosomes have been incorporated show BK channels with properties similar to those described in Table 5 and various Ca-activated K channels with more sensitivity to internal Ca, less voltage-dependence and single-channel conductances in the range of 50-100 pS in symmetrical KC1 solutions.@ All these channels are blocked by charybdotoxin.M A channel with similar properties has been described in Helix neurons.62 Its single-channel conductance is 50 pS in symmetric KC1 solutions and about 2OpS at physiological KC1 concentrations62s’26 and it is also blocked by charybdotoxin.” In squid giant axons, a Ca-activated K current blocked by external TEA and charybdotoxin has been observed (Bezanilla F., personal communication). Although the underlying single channel has not been identified, single-channel recordings do not show a channel with a conductance around 200 pS. However, the charybdotoxin-sensitive current may be mediated by a 4&50 pS channel that is observed in single-channel recordings of squid membrane.12’ The Ca sensitivity, voltage-dependence (e-fold change in conductance per 20-30 mV) and unitary conductance of these channels lie between those of the two channel types described in Table 5. Since these channels are also voltage-dependent,

RUDY

and are similar pharmacologically to BK, it may be difficult to distinguish them in macroscopic studies. Thus, 1, may be carried by all these channel subtypes. In excitable cells Ca-activated K currents are activated if Ca entry takes place during the action potential. They are responsible, in great part, for the hyperpolarizations (AHP) that follow the spike and play an important role in modulating firing frequency. There is accumulating evidence that each of the two main currents described above underlies a different component of the AHP (e.g. Refs 8, 66, 98, 111, 112, 123, 155, 166, 175, 187). Since tc is voltagedependent it tends to turn-off rapidly at voltages close to resting potential at physiological concentrations of Ca.6 It is thus associated with a fast AHP and in some neurons it even contributes to action potential repolarization (e.g. Refs 66, 111, 112, 152, 187). On the other hand, since I,,, has little voltagedependence, its decline may be more closely related to Ca diffusion away from the membrane. It is active at lower Ca concentrations and underlies the slow, long-lasting, AHP (e.g. Refs 111, 166). Many properties of slow AHPs in central neurons are similar in time course, Ca sensitivity, and resistance to TEA to those of muscle’66 and sympathetic neurons,‘07.‘55 where they are blocked by apamin. Furthermore, apamin has recently been shown to block the AHP in cat spinal motoneurqns.207 However, it has often been impossible to demonstrate apamin block of the slow AHP in brain neurons. Whether the slow AHP in these neurons is mediated by a different Ca-activated K channel or whether the insensitivity to apamin is due to problems in the drug reaching the membrane, state dependence of drug binding, or other causes remains to be established. Apamin binding to rat brain membranes and cultured neurons depends on the concentration of K ions.‘76 Variations in ion composition, for example, may be a source of apparent apamin resistance in some experiments. Apamin does bind to membranes from several parts of rat brain including the forebrain, brain stem, and the cerebellum.79” It also blocks Ca-induced Rb efflux in primary brain cultures.‘76 Furthermore, when administered intrathecally, it produces synaptic facilitation.‘02 When administered intraventricularly, it produces generalized hyperactivity which may last for more than 24 h.79 All these effects could result from blocking of the apaminsensitive channel. The two types of AHP discussed above play different roles in neuronal function. In hippocampal neurons, for example, the fast AHP helps regulate the early spike frequency while the long-lasting AHP is responsible for long-term accommodation (e.g. Refs 111, 112, 128, 129). Thus, charybdotoxin and TEA, which block the fast AHP, increase action potential duration and the frequency of the first action potentials during a depolarization.“2’87 Norepinephrine, carbachol and CAMP derivatives block the longlasting AHP without affecting the spike duration

Diversity

139

and ubiquity of K channels

or the fast AHP and prevent train adaptation.“‘~1’2~‘2*~‘3’~‘87~205 The long-lasting AHP has been associated with spike frequency adaptation in other neurons as well (e.g. Refs 11, 31,77,98,205). In addition to these Ca-activated K channels, a Ca-activated transient K current (some authors have used the term Ca-activated “A” current) has been described in some preparations. The existence of such a channel remains quite controversial. Furthermore, because it has been observed mainly in preparations where it is difficult to carry out good space-clamp and control of the internal solution, and because there is a great deal of overlap between this current and the “A” current, its analysis turns out to be rather difficult. A Ca-activated transient K channel has been proposed in bullfrog ganglion cells33~‘27 based on the sensitivity of a transient K current to Ca removal from the bath or the addition of Cd2 +, a Ca channel blocker. This current overlaps but is distinct from the “A” current. Its inactivation is faster, it is blocked by external TEA and not 4-AP, and its threshold for activation occurs at higher voltages. A current with similar properties has been reported in hippocampal CA3 cells206 In these cells, the Ca-dependent current required more positive voltages to inactivate than the “A” current. Block of a transient K current by putative Ca channel blockers or Ca removal has also been observed in barnacle motoneurons,‘60 hippocampal CA1 pyramidal cells,186Aplysin neurons,“’ and crab muscle fibers.‘” In most of these examples the current was blocked by external TEA. Ca-dependence of a transient outward current has also been observed in cardiac Purkinje fibers. 49~‘80 This current is faster than I,, and Siegelbaum and Tsien”’ have shown that it is reduced by EGTA injection as well as by Ca channel blockers. In favor of the view that there is indeed a fast transient Ca-activated current distinct from the “A” current, Salkoff”’ has described two types of transient K current in flight muscles of Drosophila. The component blocked by Ca channel blockers has a faster inactivation, higher threshold, faster recovery from inactivation and is not blocked by aminopyridines; properties that resembled the current described in bullfrog sympathetic neurons. Furthermore in several mutants of the Shaker locus, which affects the putative “A” current, the Cadependent component remains intact. Thus, in spite of the difficulties in analysis, a current with similar properties seems to be present in many cells. Furthermore, not all “A” currents are affected by preventing Ca entry, suggesting that we are dealing with a different channel. Except for its transient character, the properties of this current are similar to those carried by BK channels. Brown et ~1.‘~have suggested that this current may represent a transient component of the Ca-activated K current (I,) produced by a transient overshoot in internal Ca concentration. Further characterization is required. It remains to be established whether the current is

directly activated by Ca, and whether its kinetics are intrinsic to the channel or due to the kinetics and voltage-dependence of Ca entry. Further pharmacological studies, in particular the use of charybdotoxin, may be helpful as well. Red blood cells also contain a Ca-activated K permeability which is blocked by quinine and quinidine.‘15 Hamill** has observed two types of Ca-activated channels of 18 and 38 pS conductance in patch-clamp records of red blood cells. 5.

INWARD

RECI’IFIERS

Inward rectification is an important property of cardiac’50,‘63and skeletal muscle,’ eggs81.84and many vertebrate and invertebrate neurons (e.g. Refs 45, 105, 122, 149, 184). Table 1 described the most salient properties of inward rectifiers. A possible mechanism of inward rectification in cardiac ventricular cells has been recently proposed (see Table 1). It remains to be demonstrated whether or not this mechanism applies to inward rectifiers in other cells. Inward rectification is sufficiently distinct that less confusion exists between these channel types and the channels described before. Although no attempts at classifying them have been made, there are also several types of inward rectifiers. For example, in cardiac nodal cells there is a muscarinic activated inward rectifier channel (I*,--) which may be distinct from the inward rectifier observed in Jardiac cells which do not respond to muscarine. ‘69There are also differences in the time dependence of inward rectifiers in various cells. The inward rectifier in skeletal muscle seems to activate instantaneously and declines with time at high hyperpolarizations.” However, the one in several types of cardiac cells35,‘Nor that described in CA1 hippocampa13 and other central neurons, shows slow inward relaxations. In eggs of various animals, where inward rectification has been studied extensively, time-independent and time-dependent components have also been observed.8’,U There are also differences in the sensitivities of inward rectifiers in various cells to TEA and other channel blockers.3.‘4J83 Inward rectifiers are a very important target of modulation by neurotransmitters and second mesengers (Section 6). 6. MODULATION NEUROTRANSMITTERS

OF K CHANNELS BY AND SECOND MESSENGERS

Several neurotransmitters modify neuronal excitability by modulating the function of K channels. K channel regulation by these neurotransmitters differs from the action of neurotransmitters such as acetylcholine on nicotinic receptors. Here the channel is a separate molecule from the receptor and the effects on K channels are mediated via second messengers activated as a result of neurotransmitter-receptor interaction. As a result of such a mechanism, ion transport changes are slower than those observed for nicotinic receptor activation.

740

B. RUDY Table 6. Neurotransmitter-

Target

channel

Delayed rectifiers Slow delayed rectifier Fast delayed

rectifier

messenger-modulated

ELH

t CAMP

Inhibition

ELH

t CAMP

?

Increased rate of inactivation Phosphorylation modulates Vdependence Reduction

rectifier

?

Delayed

rectifier

Light

Cell type Aplysia bag cell neuronlsY Aplysia bag cell neuronlE9 Squid giant axon25,26 Limulus photoreceptors””

channels Repr. tract peptides

t CAMP

I K(A)

?, but see Ref. 170a Serotonin?

Ca-calmodulin kinases t kinase C

I K(A)

Norepinephrine

I WA)

ACh (muscarinic)

4W)

Ca-activated

K channels

Effect on K current

Neurotransmitter

Delayed

“A” current I K(A)

and second

(a,)

?

Inhibition’“) Incr. rate of inactivation’*” Inhibition Reduction Inhibition Inhibition

Apl.vsia bag cell neurons Hermissenda photoreceptors”“’ Hermissenda photoreceptor@’ Rat dorsal raphe nucleus cells” Rat CA1 hippocampal neurons’46

K channels Serotonin peptides Muscarine

50 PS 4WW BK

T CAMP ‘I

IAHP

ACh (must.) substance P ACh (muscarinic)

IAHP

Norepinephrine

I AHP

I K(Cal Inward rectifiers Fast I,,

Substance

P

Incr. channel opening Inhibition Reduction Inhibition

(8)

He/ix ganglion neuron@ Sympathetic neuronslSS Sympathetic neurons’53 Hippocampal pyramidal ~41s~~” Hippocampal pyramidal cell~‘~“~” Hermissenda photoreceptors’,‘70

t CAMP

Inhibition

Ca-calmodulin kinases

Inhibition

1

Inhibition Activation Enhancement

Aplvsia R 15 neuron?

Rat globus pallidus neurons’“4 Cardiac atria’b.‘ZS,,SY.‘D’

IACh

ACh (muscarinic)

I IR

Serotonin

G protein subunits t CAMP

ACh(M) substance P, others ACh (muscarinic)

not CAMP not cGMP kinase C?90” ‘1

Inhibition

Bullfrog and rat sympathetics4,5,7,8

Inhibition

Hippocampal pyramidal cells8.86

Serotonin

t CAMP

Inhibition by reducing channel opening

Aplysia sensory neurons’9,‘“’

ATP

Block

Cardiac nodal ventricular and

ATP

Block

“M” channel

“S” channel

ATP-sensitive

channels

Table 6 illustrates several examples of neurotransmitter and second messenger regulated K channels. In some of these examples the specific target of the regulation has been identified as one of the channel types described above. An example is noradrenaline modulation of an “A” current, acting through a,

atrial cells’04~‘s’ Pancreatic B-cells4’

receptors on serotonergic neurons of the dorsal raphe nucleus.‘2 It is unclear whether all the “A” current channels in these cells can be the target of modulation or if the cell contains a subtype of “A” channels capable of modification by a second messenger. Single-channel studies may help in clarifying this

Diversity and ubiquity of K channels

741

dependent and represents a distinct type of K issue. Similarly, it is unknown whether “A” currents channel. A K channel of small conductance and in all cells are capable of similar modulation. Differences between cells could depend on the existence of little voltage-dependence has been observed in guard subtypes of channels or on whether the channel is cell protoplasts of bean leaves (Viciu f~bu).“~ A K current activated by mechanical stimulation of the linked to the transducing second messenger. Studies posterior surface of Paramecia with high selectivity of the effects of second messengers on “A” channels for K over Na and blocked by external TEA also from various sources reconstituted in the same model seems distinct from the channels described above.60 A system may help to clarify this issue. As shown in light-activated K permeability with the same ionic Table 6, it is also possible for different neurotransselectivity as that of the channels described here (see mitters, acting through the same second messenger Table 1) is found in molluscan photoreceptors.73 An system, to affect the same current. Which neurotransmitter produces a given effect in a specific cell will inactivating voltage-dependent K channel has been extensively studied in T-lymphocytes.40,s’ Although depend on the presence of specific receptors. Colocalization of the receptor and the channel in a this channel is electrophysiologically very similar to reduced area of the cell may also be an important some of the voltage-dependent channels described factor, particularly for a second messenger which above, its pharmacology is very different. It is not may not diffuse or reach activating concentrations only blocked by TEA and 4-AP, but also by quinine” over the entire cell. and charybdotoxin (Cahalan M. D., personal comIn some cases, the target K channel has not been munication), which block Ca-activated K channels. identified while, in others, the target is a separate The T-lymphocyte channel, which is important for channel type and has received a specific name. One lymphocyte differentiation, is not Ca-activated. The example is the “M” channel which is inhibited by properties of this channel illustrate the difficulties in acetylcholine acting through muscarinic receptors, distinguishing between channel types even when and has been proposed to underly, in part, muscarinic using apparently more specific probes. This further excitation.4~s~7.8The “M” current is a small voltagesuggests that some relationship may exist among all dependent current. Its threshold for activation lies K channels, a point which I will take up later. at potentials negative to the resting potential. Since The lack of obvious means of distinguishing it does not inactivate, it therefore contributes to the among K channel types, and of understanding the resting K conductance of the ce11.4,8It may also play source of diversity of K channel propereties, has a role in regulating repetitive firing.4 The “S” channel caused considerable confusion in the literature. For in Aplysia neurons is a second example.39~‘09~‘79~‘8’ example, as described above, a slowly inactivating, This channel is inhibited by activation of a CAMP- highly 4-AP-sensitive K current found in hippodependent protein kinase. Increases in CAMP result campal pyramidal cells has often been described as an “A” current,76~‘65~‘86~187 but it is similar to the most from activation of a serotonin receptor. Singlechannel studies of this channel demonstrate that it frequent component of delayed rectification in the frog node of Ranvier, or to the channel in sensory has unique properties.39,‘79.‘81 It is not yet known whether channel modulation by neurons described by Stansfeld et ~1.‘~’as “novel to mammalian neurons”. This confusion is particulary kinases is the result of phosphorylation of the channel evident for “A” currents and various “delayed itself, although this is the most likely explanation. rectifier” types. The distinction is important physioThe Ca-activated K channel from Helix neurons undergoes a similar modulation in situ or when logically and pharmacologically. For example, as incorporated into model membranes.62 Since protein suggested by Stansfeld et al.‘*’ the K channel block incorporation in these membranes takes place at responsible for the convulsive actions of aminopyridines may be the slowly inactivating highly infinite dilution, it is unlikely that components not strongly associated with the channel will remain with 4-AP-sensitive current and not the “A” current, as it it in the bilayer. The modulation of K channels is usually assumed. Whatever the source of diversity, it is clear that by second messengers increases their diversity and complicates attempts to define channel types. there are distinct classes and “subclasses” of K channels, with similar properties in diverse cell types of the same animal or across species. Possible families of channels have been pointed out whenever possible. 7. SUMMARY AND PERSPECIWES We call these subclasses. However, there are clear The channels described in previous sections examples which seem to fall between categories irreillustrate the diversity and ubiquity of K channels. spective of classification scheme, particularly among However, these lists, although representative of the the voltage-dependent channels. For example, does most frequent K channels, particularly in neurons, the main K current in squid giant axons, the protoare by no means complete. For example, a K channel type “delayed rectifier”, resemble more the delayed with very large conductance has been found in the rectifiers of mammalian neurons which are blocked sarcoplasmic reticulum of skeletal and heart by external TEA and not by 4-AP or the “A” muscle.“3~‘40~‘9s This SR “maxi-K” channel is voltagecurrents which, like the squid current, are blocked by

742

B. RUDY

4-AP but not by TEA? A similar question arises if one compares activation and inactivation kinetics. It is often said that the diversity of K currents is the result of post-translational (possibly phosphorylative) modification of one or few channel types. This is based on the notion that post-translational modification of a protein produces “smaller” functional changes than changes in amino acid sequence. Many examples demonstrate that either type of change may have “small” or “large” functional consequences. At the same time, it is difficult to define which functional changes represent small modifications in protein structure and which may require larger alterations. A far better understanding of the relationship between structure and function of ion channels will be necessary to enable such judgements. The following three examples illustrate some types of structural changes that can result in functional modifications of K channels. (1) In the peptidergic bag cell neurons of Aplysia, an increse in CAMP concentration, acting presumably through channel phosphorylation, increases the rate of inactivation of a “delayed rectifier”.‘89 Here inactivation rate can become faster than “A” current inactivation in the same cells! However, the fast inactivating “delayed rectifier” found here can still be distinguished from the “A” current, because the former is present at holding potentials of -40 mV while the “A” current is already almost totally inactivated at -60 mV. (2) Coronado et aI.% have incorporated “delayed rectifiers” from lobster membranes in lipid bilayers. The single channel properties and pharmacology of these channels are very similar to those in squid giant axon (Table 1). In lipid bilayers, the single-channel conductance can change almost five times (at the same potential and ionic conditions) if the composition of the membrane is altered. The conductance increase observed in phosphatidylserine containing membranes presumably result from an increase in local K concentration, due to the larger negative surface charge. By a similar mechanism the local concentration of any cationic agent may change and thereby affect the apparent channel sensitivity to certain drugs. Ionic strength and composition also influence the surface potential and may affect single channel conductance and drug sensitivity. Lipid composition may also alter gating kinetics and voltage dependence. (3) Treatment of frog nodes of Ranvier with an amino reagent or with high pH produces a large decrease in the rate of K channel closing and reduces significantly the sensitivity to 4-AP.ls4 However, as pointed out by Hille,92 the differentiation into the various channel types described here took place tens of millions of years ago, and has been conserved in spite of many other changes. Paramecia, for example, already contain several kinds of K channels similar to the types found in vertebrates: an inward rectifier, a depolarization-activated, TEAsensitive K channel similar to “delayed rectifiers”, and several types of Ca-activated K channel (Refs 60,

110, 145, 164 and Kung C., personal communication). However, “A” currents have not been found in protozoa (Kung C., personal communication). In the eggs of coelenterates*’ one already finds the differentiation of voltage-dependent K channels with an “A” type current (see Table 4). Furthermore, similar channel types are found, often simultaneously, in different cells despite diverse active second messenger systems and probable differences in lipid composition. These findings suggest that major K channel groups represent different molecules, coded by different genes. Studies of K channel mutants in Drosophilu’9’” support this view. For example, muscles from flies carrying mutations of the Shaker gene complex lose their “A” currents or have modified “A” currents while the “delayed rectifier” remains intact.‘72.‘9’“.202 Several lines of evidence suggest that the genes involved encode structural components of the “A” channel.“‘” Recent studies of Shaker mutants also suggest that functional differences among the “A” currents may also result from different channel proteins. Solk et al. ‘*la find that in some Shaker mutants which totally eliminate I, in muscle the “A” current in cultured brain neurons, which has different kinetics and voltage-dependence (see Tables 3 and 4) remains intact. Elkins et aL6’ have described a mutation in Drosophila which specifically eliminates a calcium-dependent potassium current. Reconstitution of K channels in Xenopus oocytes by injection of messenger RNA (mRNA) from tissues rich in K channels also suggests that some of the diversity among K channels results from the presence of different channel structural components. For example oocytes injected with rat brain mRNA express various types of K channels including an “A” current and various types of “delayed rectifiers”. Selective types of K currents can be obtained after injection of size the properties fractionated mRNA.95”*‘68 Furthermore of the “A” current observed in the oocyte seem to depend on more than one mRNA species.16* At the same time it also seems valid to hypothesize that most of the channels described here evolved from a common ancestor. They all share several properties such as high selectivity for K over Na and most display the same selectivity sequence (see Table 1). Most are also blocked by Cs and Ba. As these are all very small molecules with the same charge, the channel requires a complex molecular architecture to achieve this selectivity.92 The putative precursor channel would then have evolved into several groups which represent todays major channel classes: voltage-dependent channels, Ca-activated and inward rectifiers. Each of these in turn evolved into subclasses: a few types of “I,” and “A” channels, one or a few types of inward rectifiers and two or more kinds of Ca-activated K channels. These subclasses appeared early in evolution and have since evolved independently. Protozoans already contain several types of K channels, some of which are blocked by

743

Diversity and ubiquity of K channels external TEA and by 4-AP.S3*“oWithin each subclass, gene changes resulting in new peptides, primary sequence differences in the same peptide, the addition of new subunits to a main peptide, post-translational modifications, and alterations in surrounding lipid content, ionic composition and temperature, have further resulted in different K channel types from each subclass in particular cells increasing the diversity of properties observed. Recording artifacts and problems of recording accuracy can also account for some of the “diversity” found in the literature. The reconstitution of K channels from diverse sources in the same membrane where molecular manipulations are possible, the combination of electrophysio-

logical with genetic studies in Drosophila and the isolation and characterization of messenger RNAs for various K channels now in progress in several laboratories, will help test these ideas and clarify the contributions of various parameters to K channel diversity. Acknowledgements-I

wish to thank Drs. C. Miller, M. Cahalan. F. Bezanilla and C. Kunr! for orovidina me with unpublish~ observations. I also wish to thank Drs. R. Llinas and W. Gilfy for many helpful discussions, Dr. P. Adams and his colleagues for some good ideas, Drs. C. Leonard and M. Chesler for reading the manuscript and Miss Catherine Malichio for her secretarial assistance. Supported by NIH Grant No. GM26976.

REFERENCES I.

Acos~-Urquidi J., Alcon D. L. and Neary J. T. (1984) C a’+-dependent protein kinase injection in a photor~ptor mimics biophysical effects of associative learning. Science 224, 1254-1257. 2. Adams D. J., Smith S. J. and Thompson S. H. (1980) Ionic currents in molluscan soma. A. Rev. Neurasci. 3,141-167. 3. Adams P. R. and Halliwell J. V. (1982) A hyperpolarization-induced inward current in hippocampal pyramidal cells. J. Physiol. 324, 62-633?. 4. Adams P. R., Brown D. A. and Constanti A. (1982) M-currents and other currents in bullfrog sympathetic neurons. J. Physiol. 330, 537-572. 5. Ada& P. R., Brown D. A. and Constanti A. (1982) Pharmacological inhibition of the M-current. J. Physiol. 332,

223-262. 6. Adams P. R., Constanti A., Brown D. A. and Clark R. B. (1982) Intracellular Ca2+ activates a fast voltage-sensitive K+ current in vertebrate s~pathetic neurones. Na:ure 296, 746749. 7. Adams P. R., Brown D. A. and Jones S. W. (1983) Substance P inhibits the M-current in bullfrog sympathetic neurones. Br. J. Pharmac. 79, 33C-333. 8. Adams P. R. and Galvan M. (1986) Voltage-dependent currents of vertebrate neurons and their role in membrane excitability. In Basic Mechanisms of the Epilepsies, in Advances in Neurology, Vol. 44 (eds Delgado-Escueta A., Ward A. A., Woodbury D. M. and Porter R.), pp. 137-170. Raven Press, New York. 9. Adrian R. H. (1969) Rectification in muscle membrane. Prog. Biophys. molec. Biol. 19, 34@369. 10. Adrian R. H., Chandler W. K. and Hodgkin A. L. (1970) Slow changes in potassium permeability in skeletal muscle. J. Physiol. 208, 645-668.

11. Andrew R. D. and Dudek F. E. (1984) Intrinsic inhibition in ma~~ellular

neuroendocrine cells of rat hypothal~us.

J. Physiol. 353, 171-185. 12. Aghajanian G. K. (1985) M~ulation of a transient outward current in serotonertic neurones by a,-adrenoreceptors. Nature 315, 501-503. 13. Aldrich R. W., Getting P. A. and Thompson S. H. (1978) Inactivation of delayed outward current in molluscan neuron somata. J. Physiol. 291, 507-530.

13a. Almers W. and Armstrong C. M. (1980) Survival of K permeability and gating currents in squid axons perfused with K-free media. J. gen. Physiol. 75, 61-78. 14. Armstrong C. M. (1975) Potassium pores of nerve and muscle membranes. In Membranes, a Series ofAdvances (ed. Eisenman G.). Vol. 3. DO. 325-358. Dekker. New York. 15. Armstrong C. M..and Hilie i. (1972) The inner’quaternary ammonium ion receptor in potassium channels of the node of Ranvier. J. gen. Physial. 59, 388400. 16. Armstrong C. M. and Taylor S. R. (1980) Interaction of barium ions with potassium channels in squid giant axons. Biophys. J. 3@,473-188.

17. Barish M. E. (1986) Differentiation of voltage-gated potassium current and modulation of excitability in cultured amphibian spinal neurons. J. Physiol. 375, 229-250. 18. Barrett E. F. and Barrett J. N. (1976) Separation of two voltage-sensitive potassium currents and demonstration of a tetrodotoxin-resistant calcium current in frog motoneurones. J. Physiol. US, 737-774. 19. Barrett E. F., Barrett J. N. and Grill W. E. (1980) Voltage-sensitive outward currents in cat motoneurones. J. Physio/. 304, 251-276. 20. Bartschat D. K., French R. J., Nairn A. C., Greengard P. and Krueger B. K. (1986) Cyclic AMP-dependent protein kinase modulation of single, ~cium-activate potassium channels from rat brain in planar bilayers. Neurosci. Abstr. 12, 1198. 21. Belluzzi O., Sacchi 0. and Wanke E. (1985) A fast transient outward current in the rat sympathetic neurone studied under voltage-clamp conditions. J. Physiol. 358, 91-108. 22. Belluzzi O., Sac&i 0. and Wanke E. (1985) Identification of delayed potassium and calcium currents in the rat sympathetic neurone under voltage clamp. J. Physiul. 358, 109-130. 23. Benoit E. and Dubois J. M. (1986) Toxin I from the snake Dendroaspis poiylepis polylepis: a highly specific blocker of one type of potassium channel in myelinated nerve fiber. Brain Res. 377, 374-377. 24 Benson J. A. and Levitan I. B. (1983) Serotonin increases an anomalously recitifying K+ current in the Aplysia neuron Ri5. Proc. natn. Acud. Sci. U.S.A. 80, 3522-3.525. 25. Benzanilla F. (1985) Gating of sodium and potassium channels. J. Membrane Biol. 88, 97-l 11.

744

B. RUUY

26. Benzanilla F., DiPolo R., Caputo C., Rojas H. and Torres M. E. (1985) K+ current in squid axon is modulated by ATP. Biophys. J. 47, 222a. 27. Binstock L. and Goldman L. (1967) Giant axon of Myxicola: some membrane properties as observed under voltage clamp. Science 158, 146771469. 28. Black A. R. and Dolly J. 0. (1986) Two acceptor sub-types for dendrotoxin in chick synaptic membranes distinguishable by bungarotoxin. Eur. J. B&hem. 156, 609-617. 29. Blatz A. L. and Magleby K. L. (1984) Ion conductance and selectivity of single calcium-activated potassium channels in cultured rat muscle. J. gen. Physiol. 84, I-23. 30. Blatz L. A. and Magleby K. L. (1986) Single apamin-bl~ked Ca-activated K+ channels of small conductance in cultured rat skeletal muscle. Nature 323, 718-720. 31. Bourque C. W., Randle J. C. R. and Renaud L. P. (1985) ~lcium-de~ndent potassium conductance in rat supraoptic nucleus neurosecretory neurons. .I. Neurophysiol. 54, 1375-l 382. 32. Boyett M. R. (1981) A study of the effect of the rate of stimulation on the transient outward current in sheep cardiac Purkinje fibres. J. Physiol. 319, l-22. 33. Brown D. A., Constanti A. and Adams P. R. (1982) Calcium-dependence of component transient outward current in bullfrog ganglion cells. Neurosci. Abstr. 8, 252. 34. Brown D. A., Constanti A. and Adams P. R. (1983) Ca-activated potassium current in vertebrate sympathetic neurons. Cell Calcium 4, 407420. 35. Brown H. and DiFrancesco D. (1980) Voltage-clamp investigations of membrane currents underlying pace-maker activity in rabbit ino-atria1 node. J. Ph~~~iu~. 308, 331-351. 36. Breitwieser G. E. and Szabo G. (1985) Uncoupling of cardiac muscarinic and adrenergic receptors from ion channels by a guanine nucleotide analogue. Nature 317, 538-W. 37. Burgess G. M., Claret M. and Jenkinson D. H. (1981) Effects of quinine and apamin on the Ca-dependent K permeability of mammalian hepatocytes and red cells. J. Physiol. 317, 67-90. 38. Byrne J. H. (1980) Analysis of ionic conductance mechanisms in motor cells mediating inking behavior in Aplysiu cali/ornica. J. Neurophysiol. 43, 630-650. 39. Camardo J. S., Shuster M. J., Siegelbaum S. A. and Kandel E. R. (1983) Modulation of a specific potassium channel in sensory neurons of Apiysia by serotonin and CAMP-dependent protein phosphorylation. Cold spring Harbor S,vmp. quant. Biol. 48, 213-220. 40. Cahalan M. D., Chandy K. G., DeCoursey T. E. and Gupta S. (1985) A voltage-gated potassium channel in human T-lymphocytes. J. Physiol. 358, 197.-237. 41. Chabala L. D. (1984) The kinetics of recovery and development of potassium channel inactivation in perfused squid (Loligo pealei) giant axons. J. Physiol. 356, 193.-220. 42. Connor J. A. (1975) Neural repetitive firing: a comparative study of membrane properties of crustacean walking leg axons. J. Neurophysiol. 38, 922.-932. 43. Connor J. A. and Stevens C. F. (1971) Voltage clamp studies of a transient outward membrane current in gastropod neural somata. J. Physiol. 213, 21-30. 44. Connor J. A. and Stevens C. F. (1971) Predictions of repetitive firing behavior from voltage clamp data on an isolated neurone soma. J. Physiof. 213, 31-53. 45. Constanti A. and Gaivan M. J. (1983) Fast inward-rectifying current accounts for anomalous rectification in olfactory cortex neurons. J. Physiol. 335, 153.. 178. 46. Conti F. and Neher C. (1980) Single channel recordings of K currents in squid axons. Nature 285, 14C-143. 47. Cook D, L. and Hales C. N. (1984) Intracellular ATP directly blocks K+ channels in pancreatic B cells. Nature 311, 271.-273. 48. Cooper E. and Shrier A. (1985) Single-channel analysis of fast transient potassium currents from rat nodose neurons. J. Physiol. 369, 199~-208. 49. Coraboeuf E. and Carmeliet E. (1982) Existence of two transient outward currents in sheep cardiac Purkinje fibers. Pjfiiger.7 Arch. 392, 352-359. 50. Coronado R., Latorre R. and Mautner H. G. (1984) Single potassium channels with delayed rectifier behavior from lobster axon membranes. ~iophys. J. 45, 289-299. 51. DeCoursey T. E. Chandy K. G., Gupta S. and Cahalan M. D. (1984) Voltage-gate K+ channels in human T lymphocytes: a role in mitogenesis? Nature 307, 4655468. 52. Dingledine R. (1984) Brain Slices. Plenum, New York. 53. Deitmer J. W. (1982) The effects of tetraethylammonium and other agents on the potassium mechanoreceptor current in the ciliate Stylonychia. J. exp. Biol. %, 239-249. 54. Dubinsky J. M. and Oxford G. S. (1985) Dual modulation of K channels by thyrotropin-releasing hormone in clonal pituitary cells. Proc. natn. Acad. Sci. U.S.A. 82, 42824286. 55. Dubois J. M. (1981) Evidence for the existence of three types of potassium channels in the frog Ranvier node membrane. J. Phys. 318, 297.-316. 56. Dubois J. M. (1982) Properties and physiological roles of K + currents in frog myelinated nerve tibres as revealed by 4-aminopyridine. In Advances in the Biosciences (eds Lechat P. et al.), Vol. 35. Aminop.yridines and s~m~iariyacring drugs, pp, 43-51. Pergamon Press, Oxford. 57. Dubois J. M. (1983) Potassium currents in the frog node of Ranvier. Prog. Biophys. molec. Biol. 42, 1-~20. 58. Duval A. and Leoty C. (1980) Ionic currents in slow twitch skeletal muscle in the rat. J. Physioi. 307, 23-41. 59. Duval A. and Leoty C. (1980) Comparison between the delayed outward current in slow and fast skeletal muscle in the rat. J. Physiol. 307, 43-57. 60. Eckert R. and Brehm P. (1979) Ionic mechanisms of excitation in Paramecium. A. REV. Biophys. Bioengng 8,353.--383. 61. Elkins T., Ganetzyky B. and Wu C. F. (1976) A Drosophila mutation that eliminates a calcium-dependent potassium current. Pror. natn. Acad. Sci. U.S.A. 83, 8412-8419. K+ channel activity by protein 62. Ewald D., Williams A. and Levitan I. B. (1985) Modulation of single Ca ‘+-dependent phosphorylation. Nature 315, 503-506. 63. Farley J. and Auerbach S. (1986) Protein kinase C activation induces conductance changes in ~errnj.T.senda photoreceptors like those seen in associative learning. Nature 319, 220-223.

Diversity and ubiquity of K channels

745

64. Farley J. and Rudy B (1988) Multiple types of voltage-dependent

Ca-activated K channels of large conductance in rat brain synaptosom~ membranes. B&$zJx J. In press. K+ channel in pancreatic islet cells can be 65. Findlay I., Dunne M. J. and Peterson 0. H. (1985) High~ndu~~ce activated and inactivated by internal calcium. J. Membrane Biof. 83, 169-175. 66. Fournier E. and Crepe1 F. (1984) Electrophysiological properties of dentate granule cells in mouse hippocampal slices maintained in uifro. Brain Res. 311, 75-86. 67. Frankenhaeuser B. and Huxley A. F. (1964) The action potential in the myelinated nerve fibre of Xenopus laevis as computed on the basis of voltage clamp data. J. Physiol. 171, 302-315. 68. Galvan M. and Sedlmeir C. (1984) Outward currents in voltage-clamped rat sympathetic neurones. J. Physiol. 356, 115-133. 69. Getting P. A. (1983) Mechanisms of pattern generation underlying swimming in Trifonia. III. Intrinsic and synaptic mechanisms for delayed excitation. J. ~europhysio~. 49, 10361051. 70. Giles W. R. and Van Ginneken A. C. G. (1985) A transient outward current in isolated cells from the crista terminalis of rabbit heart. J. Physial. 368, 2433264. 71. Giles W. R. and Shibata E. F. (1985) Voltage clamp of bullfrog cardiac pace-maker cells: A quantitative analysis of potassium currents. J. Physiol. 368, 265-292. 12. Gilly W. F. and Armstrong C. M. (1982) Slowing of sodium channel opening kminetics in squid axon by extracellular zinc. J. gen. Physiol. 79, 935-964. 73. Gorman A. L. F., Woolum J. C. and Cornwall M. C. (1982) Selectivity of the Ca-activated and light-dependent K channels for monovalent cations. Biophys. J. 38, 319-322. 14. Goldman L. and Schauf C. L. (1973) Quantitative description of sodium and potassium currents and computed action potentials in Myxicofa giant axons. f. gen. Physiol. 61, 361-384. 75. Gillespie J. I. and Hutter 0. F. (1975) The actions of 4-~inopy~dine on the delayed potassium current in skeletal muscle fibres. f. P&.&l. 252, 70P. 76. Gustafsson B., Galvan M., Grafe P. and Wigstrom H. (1982) Transient outward current in a mammalian central neurone blocked by 4-aminopyridine. Nature 299, 252-254. 77. Gustafsson B. and Zangger P. (1978) Effect of repetitive activation on the afterhyperpolarization in dorsal spinocerebellar tract neurones. J. Physiol. 275, 303-319. 78. Gustin M. C., Martinac B., Saimi Y., Culberston M. R. and Kung C. (1986) Ion channels in yeast. Science 233, 1195-l 197. 79. Haberman E. and Fischer K. (1978) In hreurotoxins. Tools in Neurobiology (eds Cecarelli B. and Clementi F.). Plenum Press, New York. 79a. Hab~~ann E. and Fischer K. (1979) Bee venom ~eurotoxin (apamin): Iodine labeling and characte~zation of binding sites. EUF. J. Biochem. 94, 355364. ion Channels in Cell Membrane. Phylogenetic and Devetopmenral 80. Hagiwara S. (1983) h4embrane Potential-Dependent Approaches. Raven Press, New York. Hagiwara S. and Jaffe L. A. (1979) Electrical properties of egg cell membranes. A. Rev. Biophys. Bioengng 8,385-416. tf:: Hagiwara S. and Kawa K. (1984) Calcium and potassium currents in spermatogenic cells dissociated from rat semiferous tubules. J. Physiol. 356, 135-149. 83. Hagiwara S., Kusano K. and Saito N. (1961) Membrane changes in Onchidium nerve cell in potassium-rich media. J. Physiol. 155, 470-489. 84. Hagiwara S., Miyasaki S. and Rosenthal N. P. (1976) Potassium current and the effect of cesium on this current during anomalous rectification of the egg cell membrane of a star&h. J. gen. Physioh 67, 621638. 85. Hagiwara S., Yoshida S. and Yoshii M. (1981) Transient and delayed potassium currents in the egg cell membrane of the coelenterate, Renilla koehkeri. J. Physiol. 318, 123-141. 86. Halliwell J. V. and Adams P. R. (1982) Voltage clamp analysis of muscarinic excitation in hippocampal neurons. Brain Res. 250, 71-92.

87. Halliwell J. V., Othman L. B., Pelchen-Matthews A. and Dolly J. 0. (1986) Central action of dendrotoxin selective reduction of a transient K-conductance in hippocampus and binding to localized acceptors. Proc. nam. Acad. Sri. U.S.A. 83, 493497.

88. Hamill 0. P. (1983) Potassium and chloride channels in red blood cells. In Single-channel Recording (eds Sakmann 8. and Neher E.), pp. 451-471. Plenum Press, New York. 89. Hermann A. (1985) Chary~otoxin specifically blocks Ca-activated K conductance of Aplysia neurons. Neurosci. Absrr. 11, 789. 90. Hermann A. and Hartung K. (1983) Ca activated K conductance in molluscan neurons. Cell. Calcium 4, 387-5. 90a. Higashida H. and Brown D. A. (1986) Two uolvohosnhatidvlinositide metabolites control two K currents in a neuronal cell. Nature 323, 333-335.‘ _ .I _ 91. Hille B. (1967) The selective inhibition of delayed potassium currents in nerve by tetraethylammonium ion. J. gen. Physiol. SO, 1287-1302. 92. Hille B. (1984) Ionic Channels of Excitable Membranes. Sinauer Associates, Sunderland, MA. 93. Hodgkin A. L. (1948) The local electrical changes associated with repetititve action in a nomedullated axon. J. Physiol. 107, 1655181. 94. Hodgkin A. L. and Huxley A. F. (1952) The components of membrane conductance in the giant axon of Z&go. J. Physjol. 116, 473-496. 95. Hodgkin A. L. and Huxley A. F. (1952) A quantitative description of membrane currents and its application to conduction and excitation in nerve. J. PhysioL 117, 500-544. 9Sa. Hoger J. H., Ahmed I., Davidson N., Lester H. and Rudy B. (1987) Diversity of voltage-dependent potassium channels induced in Xenopus oocytes by total and fractionated rat brain mRNA. Neurojci. Absrr. 13, 177. 96. Hugues M., Romey G., Duval D., Vincent J. P. and Lazdunski M. (1982) Apamin as a selective blocker of the Ca-dependent K channel in neuroblastoma cells. Voltage-clamp and biochemical characterization. Proc. nom. Acad. Sci. U.S.A. 79, 1308-1312. 97. Hume J. R. and Giles W. (1983) Ionic currents

153194.

in single isolated bullfrog atria1 cells. J. gen. Physiol.

81,

B. RUDY

746

98. Hotson J. R. and Prince D. A. (1980) A Ca-activated hyperpolarization follows repetitive firing in hippocampal neurons. J. Neurophysiol. 43, 409-419. 99. Jahnsen H. and Llinas R. (1984) Ionic basis for the electroresponsiveness and oscillatory properties of guinea-pi8 thalamic neurons in vitro. J. Physiol. 349, 227-247. 100. Josephson I. R., Sanchez-Chapula J. and Brown A. M. (1984) Early outward current in rat single ventricular cells, Circulation Res. 54, 157-162. 101. Junge D. (1985) Calcium dependence of A-currents in perfused Aptysia neurons. Brain Rex 346, 294-300. 102. Jurna I. and Habermann E. (1983) Intrathecai apamin selectively facilitates activity in ascending axons of rat spinal cord evoked by stimulation of afferent C fibers in sural nerve. Brain Res. 280, 186189. 103. Kaczmarek K. L., Kaczmarek L. K. and St~mwas~r R. (1984) A voltage clamp analysis of current underIying CAMP-induced membrane modulation in isolated peptidergic neurons of Apfysia. J. ~europh.ys~o~. 52, 36349. 104. Kakei M., Noma A. and Shibasaki T. (1985) Properties of adenosine-triphosphate-regulated potassium channels in guinea-pig ventricular cells. J. Physiol. 363, 441462. 105. Kandel E. R. and Taut L. (1966) Anomalous rectification in the metacerebral giant cells and its consequences for synaptic transmission. J. Physiol. 183, 2877304. 106. Kasai II., Kameyam D., Yamaguchi K. and Fukuda J. (1986) Single transient K channels in mammalian sensory neurons. Biophys. J. 49, 1243-1247. 107. Kawai T. and Watanabe M. (1986) Blockade of Ca-activated K conductance by apamin in rat sympathetic neurons. 3~. J. Pharmac. 87, 2255232. 108. Kenyon J. L. and Gibbons W. R. (1979) ~Aminopy~dine and the early outward current in sheep cardiac Purkinje fibers. J. gen. Physioi. 73, 139-157. 109. Klein M., Camardo J. and Kandel E. R. (1982) Serotonin modulates a specific potassium current in the sensory neurons that show presynaptic facilitation in Aplysia. Proc. tram. Acad. Sci. U.S.A. 79, 5713-5717. 110. Kung C. and Saimi Y. (1982) The physiological basis of taxes in Paramecium. A. Rev. Physiol. 44, 519-534. 111. Lancaster B. and Adams P. R. (1986) Calcium-dependent current generating the afterhyperpolarization of hippocampal neurons. J. Neurophysiol. 55, 126881282. 112. Lancaster B., Madison D. V. and Nicoll R. A. (1986) Charybdotoxin selectively blocks a fast Ca-dependent afterhyperpolarization (AHP) in hippocampal pyramidal cells, Neurosci. Abstr. 12, 560. 113. Latorre R. and Miller C. (1983) Conduction and selectivity in potassium channels. J. membrane Riot. 71, 11-30. 114. Latorre R., Vergara C. and Hidalgo C. (1982) Reconstitution in planar bilayers of Ca2+-dependent potassium channel from transverse tubule membranes isolated from rabbit skeletal muscle. Proc. natn. Acad. Sci. U.S.A. 79, 8055809. 115. Lew L. V. and Ferreira H. G. (1978) Calcium transport and the properties of a calcium-activated potassium channel in red cell membranes. In Current Topics in Membranes and Transport (eds Kleint-Zeller A. and Bronner F.), Vol. 10, pp. 217-277. Academic Press, New York. 116. Leonard R. J. and Lisman J. E. (1981) Light modulates voltage-dependent potassium channels in Limulus ventral photoreceptors. Science 212, 127331275. 117. Lewis R. S. and Hudspeth A. J. (1983) Voltage- and ion-dependent conductances in solitary vertebrate hair cells. Nafure 3&i, 538-541.

118. Lipkin S., Farley J. and Rudy B. (1986) Protein kinase C effects on single K channels from mammalian brain. Sot. Neurosci. Abstr. 13, 1343.

119. Lisman J., Fain G. and Swan M. (1978) Vol~ge-dependent conductance in Limulu.~ ventral photore~pto~. Biot. Bull. 135,453-454. 120. Lfano 1. and Bezanilla F. (1983) Bursting activity of potassium channels in the cut-open axon. Biophw. 1. 41, 38a. 121. Llano I. and Bezanilla F. (1985) Two types of potassium channels in the cut-open squid giant axon. Eiophys. J. 47, 22la. 122. Llinas R. (1984) Comparative electrobiology of mammalian central neurons. In Brain Slices (ed. Dingledine R.), pp. 7--24. Plenum, New York. 123. Llinas R. and Yarom Y. (1981) Electrophysiology of mammalian inferior olivary neurons in vitro. Different types of voltage-de~ndent ionic conductances. J. Physiol. 315, 549567. 124. Llinas R. and Yarom Y. (1986) Oscillatory properties of guinea-pig inferior olivary neurons and their pharma~logi~l modulation: an in vitro study. J. Physioi. 376, 163-182. 125. Lo8othetis D. E., Kurachi Y., Galper J., Neer E. J. and Clapham D. E. (1987) The subunits of GTP-binding proteins activate the muscarinic K channel in heart. Nature 325, 321.-326. 126. Lux H. D., Neher E. and Marty A. (1981) Single channel activity associated with the calcium-dependent outward current in Helix pomatia. Ppiisers Arch. 389, 293-295. 127. MacDermott A. B. and Weight F. F. (1982) Action potential repolarization may involve a transient Ca2’-sensitive outward current in a vertebrate neurone. Nafure 300, 185-188. 128. Madison D. V. and Nicoll R. A. (1982) Noradrenaline blocks accommodation of pyramidal cell discharge in the hippocampus. Nature 299, 636638. 129. Madison D. V. and Nicoll R. A. (19841 Control of repetitive discharge of rat _ pyramidal neurones in uilro. J. Physiol. . 1

354, 319.-331.

130. Madison D. V. and Nicolt R. A. (1986) Actions of noradrenaline recorded intracelluiarly in rat hippocampal CAI pyramidal neurones, in vitro. J. Physiol. 372, 221-244. 131. Madison D. V. and Nicoll R. A. (1986) Cyclic adenosine 3’,5’-monophosphate mediates beta-receptor actions of

noradrenaline in rat hippocampal pyramidal cells. J. Physiol. 372, 245259. 132. Marty A. (1981) Calcium-dependent channels with large unitary conductance in chromaffin cell membranes. Nature 291, 497-500. 133. Marty A. and Neher E. (1982) Ionic channels in cultured rat pancreatic islet cells. J. Physiol. 326, 3637P.

134. Maruyama Y., Gallagher D. V. and Peterson 0. H. (1983) Voltage and Ca-activated K channel in basolateral acinar cell membranes of mammalian salivary glands. Nature 302, 8277829. 135. Matsuda H., Saigusa A. and Irisawa H. (1987) Ohmic conductance through the inwardly rectifying K channel and blocking by internal Mg. Nature 325, 156159.

Diversity and ubiquity of K channels

747

136. Meech R. W. and Strumwaser F. (1970) Intracellular calcium injection activates potassium conductance in Aplysiu nerve cells. Fedn Proc. Fedn Am. Sots exp. Biol. 29, 835. 137. Meech R. W. (1978) Calcium-dependent potassium activation in nervous tissues. A. Rev. Biophys. Bioengng 7, l-18. 138. Meech R. W. (1972) Intracellular calcium injection causes increased potassium conductance in Aplysiu nerve cells. Comp. Biochem. P/rysiol. 42A, 493-499. 139. Meves H. and Pichon Y. (1977) The effect of internal and external ~aminopy~dine one the potassium currents in intra~llularly perfused squid giant axons. J. Physiol. 268, 51 I-532. 140. Miller C. (1978) Voltage-gated cation conductance channel from fragmented sarcoplasmic reticulum. Steady-state electrical properties. J. Membrane Biol. 40, l-23. 141. Miller C., Moczydlowski E., Latorre R. and Phillips M. (1985) Charybdotoxin, a protein inhibitor of single Ca2+-activated K+ channels from mammalian skeletal muscle. Nature 313, 316318. 142. Mirolli M. (1981) Fast inward and outward current channels in a non-spiking neurone. Nature 292, 251-253. 143. Moczy~owski E. and Latorre R. (1983) Gating kinetics of Ca’+-activated potassium channels from rat muscle incorporated into planar lipid bilayers: evidence for two volta~~e~ndent Ca r+ binding reactions. f. gen. Physiol. 82, 511-542.

144 Mounier Y. and Vassort 6. (1975) Evidence for a transient potassium membrane current dependent on calcium influx in crab muscle fibre. J. Physiol. 251, 609625. 145 Naitoh Y. (1982) Protozoa. In Electrical Conductance and Behaviour in ‘Simple’ Invertebrates (ed. Shelton G. A. B.), pp. 148. Clarendon Press, Oxford. 146. Nakajima Y., Nakajima S., Leonard R. J. and Yamaguchi K. (1986) Acetylcholine raises the excitability by inhibiting the fast transient potassium current in cultured hip~ampa~ neurons. Proc. natn. Acud. Sci. U.S.A. 83, 3022-3026. 147. Neher E. (1971) Two transient current components during voltage clamp in snail neurons. J. gen. Physiot. 61,385399. 148. Neher E. and Lux H. D. (1972) Differential action of TEA on two K-current components of a molluscan neurone. Pfiiigers Arch. 336, 87-100.

149. Nelson P. G. and Frank K. (1967) Anomalous rectification in cat spinal motoneurons and effect of polarizing currents on excitatory postsynaptic potential. 1. Neurophysiof. 30, 1097-l 113. 150. Noble D. (1984) The surprising heart: A review of recent progress in cardiac electrophysiology. J. Physiol. 353, 156159.

151, Noma A. (1983) ATP regulated K+ channels in cardiac muscle. Nature 305, 147-148. 152. Obaid A. L., Langer D. and Sal&erg B. M. (1985) Charybdotoxin (CTX) selectively blocks a calcium-mediated potassium conductance that contributes to the action potential recorded optically from nerve terminals of the frog neurohypophysis. Sot. Neurosci. Abstr. 11, 789. 153. Pallotta B. S., Magleby K. L. and Barrett J. N. (1981) single channel recordings of a Ca*+ activated K+ current in rat muscle cell culture. Nature 293, 471-474. 154, Papone P. A. and Cahalan M. D. (1984) Chemical modification of potassium channels in myelinated nerve fibers. Biophys. J. 45, 62-64.

155. Pennefather P., Lancaster B., Adams P. R. and Nicoll R. A. (1985) Two distinct Ca-dependent K currents in bullfrog sympathetic ganglion cells. Proc. nam. Acud. Sci. U.S.A. 82, 3040-3044. 156. Petersen 0. H. (1984) The mechanism by which cholecystokinin peptides excite their target cells. Biosci. Rep. 4, 275-283.

157. Peterson 0. H. and Maruyama Y. (1984) Calcium-activated potassium channels and their role in secretion. Nature 307, 693696.

158. Partridge L. D. and Stevens C. F. (1976) A mechanism for spike frequency adaptation. J. Physiol. 256, 315-332. 159. Pfaffinger P. J., Martin J. M., Hunter D. D., Nathanson N. M. and Hille B. (1985) GTP binding proteins couple cardiac muscarinie receptors to a K channel. Nature 317, 536538. 160. Premack B. A., Ruben P. and Thompson S. H. (1984) Calcium-sensitive transient K current influences rhythmic motor output in barnacle neurons. Neurosci. Abstr. 10, 144. 161. Quinta-Ferreira M. E., Rojas and Arispe N. (1982) Potassium currents in the giant axon of the crab carcinus maenas. J. Membrane

Bioi. 66, 171-181.

162. Reeves R., Farley J. and Rudy B. (1986) CAMP dependent protein kinase opens several K channels from mammalian brain. Sot. Neurosci. Abstr. 13, 1343. 163. Reuter H. (1984) Ion channels in cardiac membranes. A. Rev. Physiol. 46, 473484. 164. Richard E. A., Saimi Y. and Kung C. (1986) A mutation that increases a novel calcium-activated potassium condutance of Paramecium tetraurelia. J. Membrane Bioi. 91, 173-181. 165. Rogawski M. A. (1985) The A-current: how ubiquitous a feature of excitable cells is it? Trends Neurosci. May, 214-219. 166. Romey G. and Lazdunski M. (1984) The coexistence in rat muscle cells of two distinct classes of Ca2+-dependent K+ channels with different pharmacological properties and different physiological functions. Biochem. biophys. Res. Commun. 118, 669674. 167. Rudy B. (1976) Studies on inactivation m~hanisms in nerves. Ph.D. dissertation, Cambridge University. 168. Rudy B., Hager J. H., Davidson N. and Lester H. (1987) Expression of different “A” currents in Xenopus oocytes injected with total or fractionated rat brain mRNA. Neurosci. Abstr. 13, 178. 169. Sakmann B., Noma A. and Trautwein W. (1983) Acetylcholine activation of single muscarinic K channels in isolated pacemaker cells of mammalian heart. Nature 303, 250-253. 170. Sakakibara M., Alkon D. L., DeLorenzo R., Goldenring J. R., Neary J. T. and Heldman E. (1986) Modulation of calcium-mediated inactivation of ionic currents by Car+/calmodulin-dependent protein kinase II. Biophys. J. 9, 319327. 17Oa. Sakakibara M., Collin C., Kuzirian A., Alkon D. L., Heldman E., Naito S. and Lederhendier I. (1987) Effects of a,-adrenergic agonists and antagonists on photoreceptor membrane currents. J. Neurochem. 48, 405-416. 170b. Sakakibara M., Alkon D. L., Neary J. T., Heldman E. and Gould R. (1986) Inositol triphosphate regulation of photoreceptor membrane currents. Biophys. J. SO, 797-803. 171. Salkoff L. B. (1983) Drosophila mutants reveal two components of fast outward current. Nature 302, 249-251. 172. Salkoff L. B. and Wyman R. J. (1983) Ion currents in Drosophila flight muscles. J. Physiol. 337, 687-709.

748

B. RUDY

172a. Schwindt P. C. and Crill W. E. (1981) Differential effects of TEA and cations on outward ionic currents of cat motoneurons. J. Neurophysiol. 46, ‘l-16. 173. Schroeder J. I., Hedrich R. and Fernandez J. M. (1984) Potassium-selective single channels in guard cell protoplasts of Vicia faba. Nature 312, 361-362. 174. Schwarz J. R. and Vogel W. (1971) Potassium inactivation in single myelinated nerve fibres of Xenopus laevis. Pfliigers Arch. 330, 61-73.

175. Schwindt P. C., Spain W. J., Stafstrom W. E., Chubb M. C. and Crill W. E. (1984) Neocortical neurone after-hyperpolarizatidn multiple forms and anomalous response to membrane hyperpolarization. Nemo& Absfr. 10, 871.

176. Seagar M. J., Granier C. and Courau F. (1984) Interactions of the neurotoxin apamin with a Ca-activated K channel in primary neuronal cultures. J. b&l. Gem. 259, 1491-1495. 177. Segal M. and Barker J. L. (1984) Rat hippocampal neurones in culture potassium conductances. J. Neurophysiol. 51, 1409-1433.

178. Segal M., Rogawski M. A. and Barker J. L. (1984) A transient potassium conductance regulates the excitability of cultured hippocampal and spinal neurons. J. Neurosci. 4, 604-609. 179. Shuster M. J., Carmardo J. S., Siegelbaum S. A. and Kandel E. R. (1985) Cyclic-AMP-dependent protein kinase closes the serotonin-sensitive K+ channels of A&sin sensory neurons in cell-free membrane patches. Nuture 313, 392-395. 180. Siegelbaum S. A. and Tsien R. W. (f980) Calcium-activated transient outward current in calf cardiac Purkinje fibres. f. Physiol. 299, 485506. 181. Siegelbaum S. A., Camardo J. S. and Kandel E. R. (1982) Serotonin and cyclic AMP close single Kp channels in Aplysia sensory neurones. Nature 299, 413417. 18la. Solk C. K.. Zaeotta W. N. and Aldrich R. W. (1987) Sinale-channel and genetic analvses reveal two distinct A-tvoe -. potassium chaniels in Drosophila. Science 236, ‘1094109~. 182. Standen N. B., Stanfield P. R. and Ward T. A. (1985) Properties of single potassium channels in vesicles formed from the sarcolemma of frog skeletal muscle. J. Physiol. 364, 339-358. 183. Stanfield P. R. (1983) Tetraethylammonium ions and the potassium permeability of excitable cells. Rev. Physiol. Biochem. Pharmac. 97, 147.

184. Stanfield P. R., Nakajima Y. and Yamaguchi K. (1985) Substance P raises neuronal excitability by reducing inward rectification. Nature 315, 498-501. 185. Stansfeld C. E., Marsh S. J., Halliwell J. V. and Brown D. A. (1986) Aminopyridine and dendrotoxin induce repetitive firing in rat visceral sensory neurones by blocking a slowly inactivating outward current. Neurosci. LPft. 64,209-304. 186. Storm J. F. (1986) A-current and Ca-dependent transient outward current control the initial repetitive firing in hippocampal neurons. Biophys. J. 49, 369a. 187. Storm J. F. (1987) Action potential repolarization and a fast after-hyperpolarization in rat hippocampal pyramidal cells. J. Physiol. 365, 733-759. 188. Strong J. A. (1984) Modulation potassium current kinetics in bag cell neurons of Aplysia by an activator of adenylate cyclase. J. Neurosci. 4, 272222783. 189. Strong J. A. and Kaczmarek L. K. (1985) Multiple components of delayed potassium current in peptidergic neurons of Apfysia: Modulation by an activator of adenylate cyclase. J. Neurosci. 6, 819-822. 190. Tachibana M. (1983) Ionic currents of solitary horizontal cells isolated from goldfish retina. J. Physiol. 345, 329-35 I. 191. Tanaka K., Minota S., Kuba K., Koyano K. and Abe T. (1986) Differential effects of apamin on Ca++-dependent K currents in bullfrog sympathetic ganglion cells. Neurosci. Leff. 69, 233-238. 191a. Tanouye M. A., Kamb C. A., Iverson L. E. and Salkoff K. (1987) Genetics and molecular biology of ion channels in Drosophila. A. Rev. Neurosci. 9, 255-276. 192. Thompson S. H. (1977) Three pharmacologically distinct potassium channels in molluscan neurones. J. Physiol. 265, 465-488.

193. Thompson S. H. (1982) Aminopyridine block of transient potassium current. J. gen. Physiol. 80, I-18. 194. Thompson S. H. and Aldrich R. W. (1980) Membrane potassium channels. In The Cells Surjace and Neuronal Function feds Cotman C. W., Poste G. and Nicholson G. L.), Vol. 6, pp. 4985. Elsevier/North Holland, Amsterdam. 195. Tomlins B., Williams A. J. and Montgomery R. A. P. (1984) The characteri~tion of a monovalent cation-selective channel of mammaIian cardiac muscle sarcoplasmic reticulum. J. ~embr~e Bioi. 80, 191-199. 196. Trautman A. and Marty A. (1984) Activation of Ca-dependent K channels by carbamoyl choline in rat lacrimal glands. Proc. natn. Acad. Sci. U.S.A. 81, 611615.

197. Vandenberg C. A. (1987) Inward rectification of a potassium channel in cardiac ventricular cells depends on internal magnesium ions, Proc. natn. Acad. Sci. U.S.A. 84, 2560-2564. 198. Williams J. T., North R. A., Shefner S. A., Nishi S. and Egan T. M. (1984) Membrane properties of rat locus coeruleus neurons. Neuroscience 13, 137-156. 199. Wong B. S. and Binstock L. (1980) Inhibition of potassium conductance with external tetraethylammonium ion in Myxicola giant axons. Biophys. J. 32, IO37-1040. 200. Wong B. S. and Adler M. (1986) Tetraethylammonium blockade of calcium-activated potassium channels in clonal anterior pituitary cells. Pfliigers Arch. 407, 279289. potassium channels clonal anterior pituitary 201. Wong B. S., Lecar H. and Adler M. (1982) Single ~l~iurn-de~ndent cetls. Biophys. J. 39, 3 13-3 17. 202. Wu C. F. and Haugland F. N. (1985) Voltage-clamp analysis of membrane currents in larval muscle fibers of Drosophila: Alteration of potassium currents in Shaker mutants. J. Neurosci. 5, 26262640. 203. Yatani A., Codina J., Brown A. M. and Birnbaumer L. (1987) Direct activation of mammalian atrial muscarinic potassium channels by GTP regulatory protein Gk. Science 235, 2077211. 204. Yeh J. 2. Oxford G. S., Wu C. H. and Narahashi T. (1976) Dynamics of aminopyridine block of potassium channels in squid axon membrane. J. gen. fhysiol. 68, 519-539. preganglionic neuron spike and 205. Yoshimura M.. Poiosa C. and Nishi S. (1986) Noradrenaline modifies sympathetic . _ _ - afte~otential. &rain Res. 342, 370-374. 206. Zbicz K. L. and Weight F. F. (1985) Transient voltage and cal~um-de~ndent outward currents in hippocampal CA3 pyramidal neurons. J. Neurophysjoi. 53, 1038-1058.

Diversity and ubiquity of K channels 207. Zhang L. and Kmjevic K. (1987) Apamin depresses selectively the after-hyperpolarization Neurosci.

749 of cat spinal motoneurons.

Lett. 74. 5842. (Accepted 26 September

1987)

Note added in proof--The following developments since this review was prepared and submitted (February, 1987) are important to the theme and conclusions made here: 1. Apamin block of Ca-activated K channels has now been observed in neurons of at least two mammalian brain areas: in neurosecretory neurons in the rat superoptic nucleus of the hypothalamus [Bourque C. W. and Brown D. A. (1987) Neurosci. Z&t. 82, 1851 and in neurons in the guinea-pig mammillary bodies (Alonso A., personal communication). In hippocampal neurons, a Ca-activated K channel which may be responsible for the slow AHP has been identified in single channel recordings. However, this channel has unique properties and it is not blocked by Apamin. This is another example that suggests that more than one subclass of Ca-activated K channel may mediate slow AHPs. Therefore, the term &,, channel used in Table 5 for the Apamin-sensitive channel may be incorrect. 2. cDNAs for the “high threshold” “ A” channel encoded in the Shaker gene locus in Drosophila have been cloned [Kamb A., Iverson L. E. and Tanouye M. A. (1987) Cell 50,405; Papazian D. M., Schwarz T. L., Tempel B. L., Jan Y. N. and Jan L. Y. (1987) Science 237, 7941. The primary transcript of this gene undergoes extensive differential splicing generating a large diversity of products (Kamb A., Tseng-Crank J. and Tanouye M. A. Cell, submitted). At least three of these express “A” currents in Xenopus oocytes (Iverson L. E., Davidson N., Lester H., Tanouye M. and Rudy B. Proc. Narn. Acad. Ski., U.S.A., submitted; Timpe L. C., Schwarz T. L., Tempel B. L., Papazian D. M. Jan Y. N. and Jan L. Y. Nature, in press), which differ in inactivation kinetics. In all cases, however, the voltage-dependence of activation and inactivation is similar among the various clones and distinct from the “low threshold’ “ A” channels (Table 3). It is possible then that the Shaker “A” channel is a subclass of “A” channels (see Tabie 4) that then diversifies into distinct types. Knowledge of the molecular ~lationship between the Shaker “A” channels, the “low-threshold’ “ A” channels (Table 3) and other voltage-dependent K channels may eventually help in establishing a hierarchy of K channel diversification into classes, subclasses and types as suggested here. 3. G protein subunits may also activate K channels directly in the CNS [Andrade R., Malenka R. A. and Nicoll R. A. (1986) Science 234, 12611. 4. A book on ion-channel modulation, with many examples of modulation of K channels was published: Kaczmareck L. K. and Levitan I. B. (1987) Neuromodulation: The Biochemical ControlofNeuronai Excitability. Oxford University Press, New York.