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Isosteric regulation of the acetylcholine receptor
7TPS - May 1987 /Vol. 81
190
lsosteric regulation of the acetylcholine receptor Jayant B. Udgaonkar and George P. Hess Acetylcholine binding to the acetylcholine recepto: initiates three important events in the initiation and regulation of the transmission of signals between nerve cells and between nerve and muscle cells: the opening of transmembrane channels; a conformational change of the receptor resulting in an inactive protein (receptor desensitization); and bindingto a regulatory site that inhibits the formation of transmembrane channels. Jayant Udgaonkar and George Hess describe the two regulatory processes mediated by acetylcholine - the interaction of acetylcholine with a recently discovered regulatory site, and receptor desensitization, which does not involve the regulatory site and entails a conformational change of the receptor from an active to an inactive form with different ligand binding properties and which cannot form transmembrane channels. The signal transmission process that occurs at billions of synapses in the brain and between nerve and muscle cells is central to brain function. Chemical reactions at the junctions between cells are involved in signal transmission (see Fig. l), and understanding how these reactions are regulated is one of the important challenges in modem biology. More than twentv proteins and forty chemical signals have been implicated so far in these reactions. Newly developed experimental approaches, using the techniques of structural analysis, kinetics and molecular biology, have made possible dramatic advances in the study of one of these proteins, the nicotinic acetylcholine receptor1-5 in the electric organ of E. electricus and Torpedo spp. and have provided methods that can be used in studies of other receptor proteins. The regulatory mechanisms described are based on investigations of ticese two proteins. Regulation of signal transmission by AChR A popular view likens the acetylcholine receptor (AChR) to key enzymes that regulate the hundreds of biochemical reactions that occur within a cell’. A key feature of these muitisubunit regulatory enzymes is that they exist, relatively speaking, in active and jant Udgaonkar was a graduate student and George Hess is Professor of Biochemistry in the Section of Biochemisty, Molecular and Cell Biology, Di-oision of Biological Sciences, 270 Clark Hail, Cornell University, !thnca, NY 14853, USA. @I 1987, q swier Publications, Cambridge
conformations. An inactive important aspect of the regulatory mechanisms mediated by these enzymes is that compounds made by diverse biochemical reactions and which are not related in structure to the substrate of the enzyme (allosteric effecters), can bind preferentially to the active or inactive form of the enzyme, thereby increasing or decreasing its activity and integrating its fUnCtiOii with other bnocnemlcd reactions in the cell. The acetylcholine receptor, like regulatory enzymes, is a multisubunit protein’*6inthemembrane of nerve and muscle cells that can exist in .an active form that can form and transmembrane channels inactive forms that cannot (see Fig. 1). On binding of acetylcholine to the channel-activating sites of the active receptor form, a transmembrane channel . formed. The resulting flux z inorganic cations through such channels initiates an electrical signal, which is a prerequisite for signal transmission. The same binding process leads to desensitization, a slow (compared to channel opening) conversion of the active receptor protein to an inactive (non-channel-forming) receptor state7*12,13. A different inhibition process has recently been discovered in which an acetylcholine molecule binds to an inhibitory site that is different from the sites leading to channel opening and desensitizations”, and the site for nonendogenous inhibitor#‘*‘s, as indicated in Fig. 1. Binding to the
0165 - 6147l07/!$52.
inhibitory site is dependent on the transmembrane voltage and inhibition is fast compared to desensitization’,‘, but comparable in speed to channel opening. This inhibition process prevents the formation of transmembrane channels but does not affect the rate of receptor desensitization. We shall call this inhibitory site of the acetylcholine receptor, to which acetylcholine and its analogues bind, an isosteric site and we shall refer to the inhibitory process controlled by this site as isosteric regulation, to differentiate it from allosteric regulation of regulatory enzymes. We shall show how the voltage-dependent isosteric inhibition of the receptor determine the fraction of receptor molecules in the membrane that can form transmembrane channels and thereby make the signal transmission process sensitive to small fluctuations in both the transmembrane voltage and the concentration of acetylcholine. Desensitization, a much slower process involving a conformational change of the protein, particularly in combination with isosteric inhibition, can also be important in the regulation of receptor function. Isostericvoltage-dependeit inhibition of AChR The existence of a voltagedependent inhibitory site on the acetylcholine receptor, to which compounds including cationic local anesthetics bind, has been well documented in many types of The intriguing experimentP. question that arose is whether the site for cationic inhibitors is really a regulatory site for acetylcholine. Rapid chemical kinetic techniques, previously used to study the kinetic properties of the receptor before desensitization occurs2*13, were also used to answer this questionalO. The discovery of a natural and powerful inhibitor of receptor function, acetylcholine itself, which is thus an isosteric inhibitor, resulted from these studies’,‘. Using receptor preparations from the electric organs of two fish, Electrophorus electricus or Torpedo caiifornica, it was possible to determine KR, the dissociation constant of acetylcholine binding to the regulatory site, and the effect of transmembrane voltage on Ka; it was then demonstrated that this regulatory site is different
TIPS -May
1987
[Vol. 81
from the acetylcholine binding sites that control the opening of the receptor channel and desensitization and from the sites to which cationic inhibitors bindlo. The voltage-dependent inhibition of the receptor by acetylcholine has many features that make it attractive as an important regulatory mechanism: l Acetylcholine is known to be present during the transmission process at concentrations of 100 PM to 500 ~IW,which are about equal to the value of the dissociation constant of the channelactivating site but are much larger than the value of KR at the average membrane potential of electroplax cells (-70 mv). This has two important consequences: (1) The density of receptors that can form transmembrane channels is reduced to a critical value for signal transmission. The estimated density of receptors at the synapse is - 20000 per vrn*. From a determination of the effect of transmembrane voltage on KR (Refs 8, 9) we calculate that at -70 mV only 100-200 receptors per p* can still form channels. The critical number of receptors in the open-channel form per pm2 l..-Snt,ir~~ $.-W‘L6L_C .G_%_? f-_?-_l--_:2&.~~ 2=-.~=~_;-:’1s; _______ -_______ has been estimated to be 50400 per pm (Ref. 17). (2) The fraction of the receptors present that can form transmembrane channels, F, is given by? F = F&(Ka
+ Q-l
where F0 represents the fraction of receptors in the open-channel form in the absence of isosteric inhibition and L represents the acetylcholine concentration. It can be seen from the equation that when the acetylcholine concentration is much larger than Ka, changes in transmembrane voltage that affect I& have a maximum effect on F. At a transmembrane voltage of -70 mV, a change of +5 mV can increase or decrease the number of receptors capable &forming transmembrane channels by a factor of two (Ref. 9). The use of a combination of singlechannel current and rapid chemical kinetic measurements indicate&14-*6 that on exposure of the receptor to acetylcholine, isosteric regulation occurs on the same time scale as signal transmission. Cationic inhibitors can bind to the inhibitory site before the channel
191 Receptor forms involved in channel opening
lsosterically inhibited receptor form
Desensitized inactive receptor form Desensitizetion
(Conformational
change
Open-channel receptor form allowing transmembrane movement of cations IM+I Fig. 1. Schematic representation of regu/atory mechanisms at synaptic acetylcho!ine receptor. The receptor confains five subunits with a sroichomery a&6. The two acetylcholine binding sites con&o\ling channel opening and desensifkation (0) are located on the two cu-subunit& me location of the regulatorybindingsite(a) responsible for isosteric inhibition is not yet known. At the synapse, acefyfcholine (1:) teleasedby one cell in response to an e/ecftica/ siQna/ binds to the acetylcholine mceplor in fhe membrane of fhe postsynaptic cell. Before the signal can be Partsmiffed, acefyk3oljne binds to fhe regulatory& indicafed by A and characterized by thedissociition constant I&, which depends on the transmembrane &age. Winding of ace~lcholine to the sires confrolling channel opening and desensitization (0) results in the rapid (-1 ms) formatio,n of fransn--?--r--~& pnnuw-idn l-l* “,m.,.CzY,*...I*.-.Y, __.*-_.i._. OFthe receptor to its desensifized Form, indicated by Ihe square, involves a proteln conForma&nal change. The exchange of cations M+. through fhe open receptorchannelresults in an eh3ctricalsi~nal. when the amplitude of this signal reaches a critical value in a muscle cell membrane, the mUstYeWill contract, or, in a nerve ceilmembrane, the e!ectricalsignalispmpaga89din an all-Or-ItOne process to a nerve terminal at another synapse.
opens I’. Isosteric inhibition can thus determine the number of receptors that can form transmembrane channels and, therefore, regulate signal transmission. l The protein is located at critica! synapses in which messages from the nervous system are transmitted to muscles to initiate contraction. Isosteric inhibition of acetylcholine receptors in muscle appears to be a general phenomenon. It accounts for the observation that the current that initiates signal transmission at the synapse corresponds to the current flowing
through only a small fraction of the acetylcholine receptors that are actually present in the synapse” and also for the blocking of receptor channels in BC$Ills and frog muscle cell?. o Mechanisms for the short-term modulation of the transmembrane voltage of nerve cells are well known and short-term mndulation of muscle cell membranes has recently been reported”. Protein phosphorylation, recognized as important in brain function”, may be important in long-term modulation of transmembrane vo!-
TIPS - May 1987 [Vol. 81
192 tage by modifying the activity of potassium channels Wdgaonkar, unpublished). Receptordesensitization Electrophysiological measurements indicated that desensitization of the acetylcholine receptor, in contrast to the rapid inhibition involving the regulatory site occurs in the second-to-minute time scale’*ll. Desensitization was, therefore, considered to be irrelevant to receptor-mediated signal transmission, which occurs in milliseconds. A major limitation of electrophysiological experiments with cells is the inability to make measurements wi*& a few milliseconds after addition of acetylcholine, before the receptor is desensitized. Rapid chemical kinetic investigation&l3 of receptor function overcame this difficulty and led to *&e discovery of a desensitization process in E. electricks and T. caZifornicureceptors that occurs in the 100-300 ms time scale2*U,13. This process is about 100 times slower than is isosteric inhibition, does not involve the isosteric regulatory site and is accompanied by changes in ligand binding properties of the receptor, which suggests that a conformational change is involved’2*13. A much slower desensitization process, reminiscent of the one previously reported’, is also observed. Howcanthesepropertiesbeused to modulate signal transmission? Taking the electroplax receptor of E. electricus and Torpedo as a model, it can be seen that, at the transmembrane voltages typical of these cells, the binding of acetylcholine to the regulatory site reduces the effective receptor concentration to a value critical for signal transmission and makes the reactions at the synapse particuIarli sensitive to variations in the amount of acetylcholine released by the presynaptic cell. Variations in the amount of acetylcholine released have been observed at certain synapses in the sea snai120 in response to external stimuli and these variations affect the response of the animal. Desensitization may have an important regulatory function when the concentration of active receptors becomes critical for signal transmission. Although only a fati percent of the receptors may be
desensitized in the one millisecond time period during signal transmission, this may be sufficient to block a subsequent signal transmission process because recovery from desensitization is about twenty times slower than desensitization i tself. Specific phosphorylation of the acetylcholine receptor by a cAMP-dependent protein kinase increases the desensitization rate=, this may have an important effect on the regulatory mechanism -. discussed. 0
cl
0
Voltage-dependent inhibition by acetylcholine of a large fraction of the receptors in the postsynaptic cell occurs before signal transmission can occur at a synapse. The remaining effective concentration of receptor5 may be critical in determining whether a signal is transmitted. Variations in the transmembrane voltage, in conjunction with changes in the amount of acetylcholine released and the desensitization reaction, can therefore modulate transmission. Changes in both the resting potential of cells and the amount of acetylcholine released have been reported. Chemical kinetic measurements, employing rapid reaction techniques, have been as useful in elucidating the regulatory properties of the acetylcholine receptor as they were previously in elucidating the mechanisms of regulatory enzyme523 and as they are likely to be in future investigations of other receptors. Investigations of the regulatory properties of the proteins at the synapse and of the utilization of these properties in an animal’s response to external stimuli, are likely to produce exciting results with farreaching implications for our
understanding of the way the brain functions. References 1 Changeux, J.-P., Devillers-ThiBry, A. and Chemouilli, P. (1984) Science 225, 1335-lk%5 2 Hess, G. P., Cash, D. J. and Aoshima, H. (1983) Annu. Rev. Biophys. Bioeng. 12, -73 3 Stroud, R. M. and Finer-Moore, J. (1985) Annu. Rev. Cell Biol. 1,36%401 4 Brisson, A. and Unwin, P. N. T. (1985) Nature 315.474-477 5 Claudio, T. (1986) Trends Pharmacol. Sci. 7,308-312 6 Karlin, A., Cox, R., Kaldany, R.-R., Label, P. and Holtzman, E. (1983) Cold Spring Harbor Symp. Quant. Biol. 48, 1-8 7 Katz, 8. and Thesleff, S. (1957) J. Physiol. (London) 138,63-80 8 Takeyasu, K., Udgaonkar, J. B. and Hess, G. P. (1983) Biochemistry 22,59735978 9 Takeyasu, K., Shiono, S., Udgaonkar, J. B., Fujita, N. and Hess, G. P. (1986) Biochemisfy 25, 177C-1776 10 .Shiono, S., TakeyasG, K., Udgaonkar, J. B., Delcour, A. H., Fujita, N. and Hess, G. P. (1984) Biochemistry 23,68896893 11 Adams, P. R. (1981) J, Membr. Biol. 58, 161-174 12 Hess, G. P., Lipkowitz, S. and Struve, G. E. (1978) Proc. N&l Acad. Sci. USA 75,
i7oG707.
13 Hess, G. P., Cash, D. J. and Aoshima, H. (19791 Nature 282,329-331 14 ~ak&nn, B., Pahack, J. and Neher, E. (1980) Nature 286, 71-73 15 sine,.S. M. and Steinbach, J. H. (1984) Biophys. ].46,277-284 16 Ogden, D. C. and Colquhoun, D. (1985) Pmt. R. Sot. Land. 8225.329-355 17 Junge, D. (1981) Nerue and Muscle Excitation. Sinauer 18 Karpen, i. W. and Hess, G. P. (1986) Biochemisty 25,1777-1785 19 Thesleff, S. and Molgo, J. (1984) Neuroscience 9, l-8 20 Schwartz, J. H., Bemier, i., Castellucci, V. F., Palazzolo, M., Saitoh, T., Stapleton, A. and Kandel, E. R. (1983) Cold Spring Harbor Symp. Quant. Biol. 48, 811-819 21 Greengard, P. (1978) Science 199, 14& 152 22 Huganir, R. L., Delcour, A. H., Greengard, P. and Hess, G. P. (1986) Nature 321,774 23 Hammes, G. G. (1982) Enzyme Catalysis and Regulation, Academic Press
-Subscribers!
Moving house? Moving job? Please send your change of address to our Amsterdamsubscriptiondepartment: Ste!laFenley, Elsevier Science Publishers, TrendsJournalsDepartment,PO Box 2 11,lOOO AE, Amsterdam,The ktherknds.
190
lsosteric regulation of the acetylcholine receptor Jayant B. Udgaonkar and George P. Hess Acetylcholine binding to the acetylcholine recepto: initiates three important events in the initiation and regulation of the transmission of signals between nerve cells and between nerve and muscle cells: the opening of transmembrane channels; a conformational change of the receptor resulting in an inactive protein (receptor desensitization); and bindingto a regulatory site that inhibits the formation of transmembrane channels. Jayant Udgaonkar and George Hess describe the two regulatory processes mediated by acetylcholine - the interaction of acetylcholine with a recently discovered regulatory site, and receptor desensitization, which does not involve the regulatory site and entails a conformational change of the receptor from an active to an inactive form with different ligand binding properties and which cannot form transmembrane channels. The signal transmission process that occurs at billions of synapses in the brain and between nerve and muscle cells is central to brain function. Chemical reactions at the junctions between cells are involved in signal transmission (see Fig. l), and understanding how these reactions are regulated is one of the important challenges in modem biology. More than twentv proteins and forty chemical signals have been implicated so far in these reactions. Newly developed experimental approaches, using the techniques of structural analysis, kinetics and molecular biology, have made possible dramatic advances in the study of one of these proteins, the nicotinic acetylcholine receptor1-5 in the electric organ of E. electricus and Torpedo spp. and have provided methods that can be used in studies of other receptor proteins. The regulatory mechanisms described are based on investigations of ticese two proteins. Regulation of signal transmission by AChR A popular view likens the acetylcholine receptor (AChR) to key enzymes that regulate the hundreds of biochemical reactions that occur within a cell’. A key feature of these muitisubunit regulatory enzymes is that they exist, relatively speaking, in active and jant Udgaonkar was a graduate student and George Hess is Professor of Biochemistry in the Section of Biochemisty, Molecular and Cell Biology, Di-oision of Biological Sciences, 270 Clark Hail, Cornell University, !thnca, NY 14853, USA. @I 1987, q swier Publications, Cambridge
conformations. An inactive important aspect of the regulatory mechanisms mediated by these enzymes is that compounds made by diverse biochemical reactions and which are not related in structure to the substrate of the enzyme (allosteric effecters), can bind preferentially to the active or inactive form of the enzyme, thereby increasing or decreasing its activity and integrating its fUnCtiOii with other bnocnemlcd reactions in the cell. The acetylcholine receptor, like regulatory enzymes, is a multisubunit protein’*6inthemembrane of nerve and muscle cells that can exist in .an active form that can form and transmembrane channels inactive forms that cannot (see Fig. 1). On binding of acetylcholine to the channel-activating sites of the active receptor form, a transmembrane channel . formed. The resulting flux z inorganic cations through such channels initiates an electrical signal, which is a prerequisite for signal transmission. The same binding process leads to desensitization, a slow (compared to channel opening) conversion of the active receptor protein to an inactive (non-channel-forming) receptor state7*12,13. A different inhibition process has recently been discovered in which an acetylcholine molecule binds to an inhibitory site that is different from the sites leading to channel opening and desensitizations”, and the site for nonendogenous inhibitor#‘*‘s, as indicated in Fig. 1. Binding to the
0165 - 6147l07/!$52.
inhibitory site is dependent on the transmembrane voltage and inhibition is fast compared to desensitization’,‘, but comparable in speed to channel opening. This inhibition process prevents the formation of transmembrane channels but does not affect the rate of receptor desensitization. We shall call this inhibitory site of the acetylcholine receptor, to which acetylcholine and its analogues bind, an isosteric site and we shall refer to the inhibitory process controlled by this site as isosteric regulation, to differentiate it from allosteric regulation of regulatory enzymes. We shall show how the voltage-dependent isosteric inhibition of the receptor determine the fraction of receptor molecules in the membrane that can form transmembrane channels and thereby make the signal transmission process sensitive to small fluctuations in both the transmembrane voltage and the concentration of acetylcholine. Desensitization, a much slower process involving a conformational change of the protein, particularly in combination with isosteric inhibition, can also be important in the regulation of receptor function. Isostericvoltage-dependeit inhibition of AChR The existence of a voltagedependent inhibitory site on the acetylcholine receptor, to which compounds including cationic local anesthetics bind, has been well documented in many types of The intriguing experimentP. question that arose is whether the site for cationic inhibitors is really a regulatory site for acetylcholine. Rapid chemical kinetic techniques, previously used to study the kinetic properties of the receptor before desensitization occurs2*13, were also used to answer this questionalO. The discovery of a natural and powerful inhibitor of receptor function, acetylcholine itself, which is thus an isosteric inhibitor, resulted from these studies’,‘. Using receptor preparations from the electric organs of two fish, Electrophorus electricus or Torpedo caiifornica, it was possible to determine KR, the dissociation constant of acetylcholine binding to the regulatory site, and the effect of transmembrane voltage on Ka; it was then demonstrated that this regulatory site is different
TIPS -May
1987
[Vol. 81
from the acetylcholine binding sites that control the opening of the receptor channel and desensitization and from the sites to which cationic inhibitors bindlo. The voltage-dependent inhibition of the receptor by acetylcholine has many features that make it attractive as an important regulatory mechanism: l Acetylcholine is known to be present during the transmission process at concentrations of 100 PM to 500 ~IW,which are about equal to the value of the dissociation constant of the channelactivating site but are much larger than the value of KR at the average membrane potential of electroplax cells (-70 mv). This has two important consequences: (1) The density of receptors that can form transmembrane channels is reduced to a critical value for signal transmission. The estimated density of receptors at the synapse is - 20000 per vrn*. From a determination of the effect of transmembrane voltage on KR (Refs 8, 9) we calculate that at -70 mV only 100-200 receptors per p* can still form channels. The critical number of receptors in the open-channel form per pm2 l..-Snt,ir~~ $.-W‘L6L_C .G_%_? f-_?-_l--_:2&.~~ 2=-.~=~_;-:’1s; _______ -_______ has been estimated to be 50400 per pm (Ref. 17). (2) The fraction of the receptors present that can form transmembrane channels, F, is given by? F = F&(Ka
+ Q-l
where F0 represents the fraction of receptors in the open-channel form in the absence of isosteric inhibition and L represents the acetylcholine concentration. It can be seen from the equation that when the acetylcholine concentration is much larger than Ka, changes in transmembrane voltage that affect I& have a maximum effect on F. At a transmembrane voltage of -70 mV, a change of +5 mV can increase or decrease the number of receptors capable &forming transmembrane channels by a factor of two (Ref. 9). The use of a combination of singlechannel current and rapid chemical kinetic measurements indicate&14-*6 that on exposure of the receptor to acetylcholine, isosteric regulation occurs on the same time scale as signal transmission. Cationic inhibitors can bind to the inhibitory site before the channel
191 Receptor forms involved in channel opening
lsosterically inhibited receptor form
Desensitized inactive receptor form Desensitizetion
(Conformational
change
Open-channel receptor form allowing transmembrane movement of cations IM+I Fig. 1. Schematic representation of regu/atory mechanisms at synaptic acetylcho!ine receptor. The receptor confains five subunits with a sroichomery a&6. The two acetylcholine binding sites con&o\ling channel opening and desensifkation (0) are located on the two cu-subunit& me location of the regulatorybindingsite(a) responsible for isosteric inhibition is not yet known. At the synapse, acefyfcholine (1:) teleasedby one cell in response to an e/ecftica/ siQna/ binds to the acetylcholine mceplor in fhe membrane of fhe postsynaptic cell. Before the signal can be Partsmiffed, acefyk3oljne binds to fhe regulatory& indicafed by A and characterized by thedissociition constant I&, which depends on the transmembrane &age. Winding of ace~lcholine to the sires confrolling channel opening and desensitization (0) results in the rapid (-1 ms) formatio,n of fransn--?--r--~& pnnuw-idn l-l* “,m.,.CzY,*...I*.-.Y, __.*-_.i._. OFthe receptor to its desensifized Form, indicated by Ihe square, involves a proteln conForma&nal change. The exchange of cations M+. through fhe open receptorchannelresults in an eh3ctricalsi~nal. when the amplitude of this signal reaches a critical value in a muscle cell membrane, the mUstYeWill contract, or, in a nerve ceilmembrane, the e!ectricalsignalispmpaga89din an all-Or-ItOne process to a nerve terminal at another synapse.
opens I’. Isosteric inhibition can thus determine the number of receptors that can form transmembrane channels and, therefore, regulate signal transmission. l The protein is located at critica! synapses in which messages from the nervous system are transmitted to muscles to initiate contraction. Isosteric inhibition of acetylcholine receptors in muscle appears to be a general phenomenon. It accounts for the observation that the current that initiates signal transmission at the synapse corresponds to the current flowing
through only a small fraction of the acetylcholine receptors that are actually present in the synapse” and also for the blocking of receptor channels in BC$Ills and frog muscle cell?. o Mechanisms for the short-term modulation of the transmembrane voltage of nerve cells are well known and short-term mndulation of muscle cell membranes has recently been reported”. Protein phosphorylation, recognized as important in brain function”, may be important in long-term modulation of transmembrane vo!-
TIPS - May 1987 [Vol. 81
192 tage by modifying the activity of potassium channels Wdgaonkar, unpublished). Receptordesensitization Electrophysiological measurements indicated that desensitization of the acetylcholine receptor, in contrast to the rapid inhibition involving the regulatory site occurs in the second-to-minute time scale’*ll. Desensitization was, therefore, considered to be irrelevant to receptor-mediated signal transmission, which occurs in milliseconds. A major limitation of electrophysiological experiments with cells is the inability to make measurements wi*& a few milliseconds after addition of acetylcholine, before the receptor is desensitized. Rapid chemical kinetic investigation&l3 of receptor function overcame this difficulty and led to *&e discovery of a desensitization process in E. electricks and T. caZifornicureceptors that occurs in the 100-300 ms time scale2*U,13. This process is about 100 times slower than is isosteric inhibition, does not involve the isosteric regulatory site and is accompanied by changes in ligand binding properties of the receptor, which suggests that a conformational change is involved’2*13. A much slower desensitization process, reminiscent of the one previously reported’, is also observed. Howcanthesepropertiesbeused to modulate signal transmission? Taking the electroplax receptor of E. electricus and Torpedo as a model, it can be seen that, at the transmembrane voltages typical of these cells, the binding of acetylcholine to the regulatory site reduces the effective receptor concentration to a value critical for signal transmission and makes the reactions at the synapse particuIarli sensitive to variations in the amount of acetylcholine released by the presynaptic cell. Variations in the amount of acetylcholine released have been observed at certain synapses in the sea snai120 in response to external stimuli and these variations affect the response of the animal. Desensitization may have an important regulatory function when the concentration of active receptors becomes critical for signal transmission. Although only a fati percent of the receptors may be
desensitized in the one millisecond time period during signal transmission, this may be sufficient to block a subsequent signal transmission process because recovery from desensitization is about twenty times slower than desensitization i tself. Specific phosphorylation of the acetylcholine receptor by a cAMP-dependent protein kinase increases the desensitization rate=, this may have an important effect on the regulatory mechanism -. discussed. 0
cl
0
Voltage-dependent inhibition by acetylcholine of a large fraction of the receptors in the postsynaptic cell occurs before signal transmission can occur at a synapse. The remaining effective concentration of receptor5 may be critical in determining whether a signal is transmitted. Variations in the transmembrane voltage, in conjunction with changes in the amount of acetylcholine released and the desensitization reaction, can therefore modulate transmission. Changes in both the resting potential of cells and the amount of acetylcholine released have been reported. Chemical kinetic measurements, employing rapid reaction techniques, have been as useful in elucidating the regulatory properties of the acetylcholine receptor as they were previously in elucidating the mechanisms of regulatory enzyme523 and as they are likely to be in future investigations of other receptors. Investigations of the regulatory properties of the proteins at the synapse and of the utilization of these properties in an animal’s response to external stimuli, are likely to produce exciting results with farreaching implications for our
understanding of the way the brain functions. References 1 Changeux, J.-P., Devillers-ThiBry, A. and Chemouilli, P. (1984) Science 225, 1335-lk%5 2 Hess, G. P., Cash, D. J. and Aoshima, H. (1983) Annu. Rev. Biophys. Bioeng. 12, -73 3 Stroud, R. M. and Finer-Moore, J. (1985) Annu. Rev. Cell Biol. 1,36%401 4 Brisson, A. and Unwin, P. N. T. (1985) Nature 315.474-477 5 Claudio, T. (1986) Trends Pharmacol. Sci. 7,308-312 6 Karlin, A., Cox, R., Kaldany, R.-R., Label, P. and Holtzman, E. (1983) Cold Spring Harbor Symp. Quant. Biol. 48, 1-8 7 Katz, 8. and Thesleff, S. (1957) J. Physiol. (London) 138,63-80 8 Takeyasu, K., Udgaonkar, J. B. and Hess, G. P. (1983) Biochemistry 22,59735978 9 Takeyasu, K., Shiono, S., Udgaonkar, J. B., Fujita, N. and Hess, G. P. (1986) Biochemisfy 25, 177C-1776 10 .Shiono, S., TakeyasG, K., Udgaonkar, J. B., Delcour, A. H., Fujita, N. and Hess, G. P. (1984) Biochemistry 23,68896893 11 Adams, P. R. (1981) J, Membr. Biol. 58, 161-174 12 Hess, G. P., Lipkowitz, S. and Struve, G. E. (1978) Proc. N&l Acad. Sci. USA 75,
i7oG707.
13 Hess, G. P., Cash, D. J. and Aoshima, H. (19791 Nature 282,329-331 14 ~ak&nn, B., Pahack, J. and Neher, E. (1980) Nature 286, 71-73 15 sine,.S. M. and Steinbach, J. H. (1984) Biophys. ].46,277-284 16 Ogden, D. C. and Colquhoun, D. (1985) Pmt. R. Sot. Land. 8225.329-355 17 Junge, D. (1981) Nerue and Muscle Excitation. Sinauer 18 Karpen, i. W. and Hess, G. P. (1986) Biochemisty 25,1777-1785 19 Thesleff, S. and Molgo, J. (1984) Neuroscience 9, l-8 20 Schwartz, J. H., Bemier, i., Castellucci, V. F., Palazzolo, M., Saitoh, T., Stapleton, A. and Kandel, E. R. (1983) Cold Spring Harbor Symp. Quant. Biol. 48, 811-819 21 Greengard, P. (1978) Science 199, 14& 152 22 Huganir, R. L., Delcour, A. H., Greengard, P. and Hess, G. P. (1986) Nature 321,774 23 Hammes, G. G. (1982) Enzyme Catalysis and Regulation, Academic Press
-Subscribers!
Moving house? Moving job? Please send your change of address to our Amsterdamsubscriptiondepartment: Ste!laFenley, Elsevier Science Publishers, TrendsJournalsDepartment,PO Box 2 11,lOOO AE, Amsterdam,The ktherknds.