Developmental changes of the expression of voltage-dependent K+ channels in cerebellar granule cells

s59 SPLICING VARIANTS OF KIR3.2 ARE ASSEMBLED BUT PLAY DISTINCT ROLES IN THE G PROTEIN-GATED INWARDLY RECTIFYING POTASSIUM CHANNEL IN RAT SUBSTANTIA NIGRA

003

ATSUSHI INANOBE’, HIROSHI HIBINO’, SHIGETO MATSUMOTO’, YUKIKO YOSHIMOTO’, YOSHIYUKI HORIO’, KEN-ICHIRO MORISHIGE’, YOSHIMITSU TOKUNAGA’. TOSHIHIRO MAEDA2, YUTAKA HATA3, YOSHIMI TAKAI’, YOSHIHISA KURACHI’ ‘Departments of Pharmacology II and “Molecular Biology and Biochemistry, 565-0087.

?Department

Faculty of Medicine, Osaka University, Osaka

of Anatomy I, Shiga University of Medical Science, Shiga 520-2192, Takai Biotimer Project, ERATO,

Japan Science and Technology Corp., Kobe 65 l-224 1. G protein-gated K+ (K,) channels generate slow inhibitory postsynaptic

potentials (IPSPs).

Current opinion suggests that

neuronal K, channels are heterotetramers of GIRKIKir3.1 and GIRK2/Kir3.2. However biochemical and immunological analyses revealed that the K, channel in rat SN was composed of splicing variants of Kir3.2. Kir3.2a+Kir3.2c hetero- and Kir3.2c homotetramers.

and did not contain Kir3.1.

Immunohistochemical

studies indicated that Kir3.2a and Kir3.2c were co-

localized specifically at the postsynaptic membrane on the dendrites of dopaminergic bind PSD-95. a postsynaptic density anchoring protein. The K, channels composed

neurons, Kir3.2c. but not Kir3.2a. could of Kir32a + Kir3.2c. or Kir3.2a alone

expressed in Xenopus oocytes were activated by D2-dopamine receptor or G protein stimulation, but that of Kir3.2c alone was not. This study reveals that the splicing variants of Kir3.2 may play distinct roles in the control of function and subcellular localization of K, channels and their homo- or heterotetramers may involve in different mechanisms generating

heterologously

slow IPSPs in dopaminergic

Developmental Changes Cerebellar Granule Cells.

004 Riichi

Shibata,

Department Okazaki

neurons of the SN.

Yoshihiko

of Neural

Wakazono,

Information,

of

the

Expression

Kensuke

Nakahira,

National

Institute

of

Voltage-dependent

Kazuhiro

K’ Channels

in

Ikenaka

for Physiological

Sciences,

Myodaiji-thou,

444-0867

To investigate the role of voltage-dependent K’ channels in developing cerebellar granule cells. we examined temporal expression pattern of K’ channels during development. Kv4.2 protein, one of the A-type K* channels, was detected in internal granule layer (IGL) and premigratory zone (PMZ) of external granule layer (EGL) of neonatal mice as shown by in situ hybridization previously. This result shows that granule cells began to express Kv4.2 before migration, We also examined the developmental change of expression of the voltage-gated K’ channel proteins in microexplant culture of cerebellum granule cell dissected from neonatal mice EGL. At 2DIV. Kv3.1 protein was detected in parallel fiber of granule cells and weak signals of Kv4.2 protein were detected in soma of small number of granule cells. At 6DIV. Kvl.I protein was also expressed in parallel fiber at a low level in addition to Kv3.1. and the number of cells and intensity of Kv4.2 protein signal increased. Furthermore, we measured developmental change of action potential by using whole-cell patch clamp method. No or single action potential was observed in immature bipolar granule cells. On the other hand, repetitive firing was observed in mature T-shape granule cells. The relationship between change of action potential and expression of K’ channels during development will be discussed.

005 MICHIHISA

IDENTIFICATION

OF A DOMAIN

Kvl.2 POTASSIUM

CHANNELS

MIYAKE & KENTARO

Faculty of Pharmaceutical Whole-cell

cells.

dependent

facilitation.

various chimeric

Stimulation

K’ channel containing the remainder

of Kv1.5 exhibited

S2 and S3) showed

the transmembrane

segment

of Kv 1.2 K’ channels.

Hokkaido University

OF

Sapporo 060-0812

protocol

revealed

the domain of the channel

of Kvl.2

the Kvl.2

FACILITATION

IZUMI

with a double-pulse

To identify

constructs

IN VOLTAGE-DEPE.NDENT

was used to study the K’ currents through Kvl.2 channels transiently

patch clamp recording

fibroblast

(between

Sciences,

INVOLVED

and Kvl.5,

no facilitation

facilitation

transmembrane

segment

K’ current

important

no such facilitation

of K’ current. However,

of the current

S2 and cytoplasmic

which is critically

a channel showing

domain of N-terminal,

that the Kvl.2

depolarization-

for the facilitation,

S 1 and linker Ll (between

a chimera containing

important

showed

in L-929

we made

of evoked K’ current. A chimeric

like that of the parent Kv1.2 channel.

linker L2 are critically

expressed

S 1 and S2) and

Kv1.2 of N-terminal Thus, it was concluded

for depolarization-dependent

to L2 that

facilitation