Akiko Kose

Electron microscopic localization of γ-and βII-subspecies of protein kinase C in rat hippocampus

Abstract The subcellular distributions of the γ- and βII-subspecies of protein kinase C (γ- and βII-PKC) were studied in the rat hippocampus by light and electron microscopic immunocytochemistry. Both subspecies were abundant in the hippocampus with distinct subcellular distributions. The immunoreactivity of γ-PKC was observed throughout the Ammons horn, while intense βII-PKC immunoreactivity was observed predominantly in the CA1 region. γ-PKC was distributed diffusely through the cytoplasm of pyramidal cells from the perikarya to the dendritic spines. In contrast, βII-PKC was concentrated around the Golgi complex and present diffusely in distal dendrites, except for the dendritic spines. Neither PKC subspecies could be detected in the presynaptic terminal. The postsynaptic localization of γ- and βII-PKC in CA1 suggests that both PKC subspecies may correlate to long-term potentiation in the CA1 region contributing to the postsynaptic side. γ-PKC may have a specific function not only in CA1 but also in the mossy fiber-CA3 pathway at the postsynaptic side. βII-PKC may have another function concerning the Golgi complex in CA1.

Possible roles of protein kinase C in neurotransmission.

Protein kinase C (PKC) is an ubiquitous Ca2+-activated phospholipiddependent protein kinase abundant in the central nervous system(1–4). This enzyme is activated by 1,2-diacylglycerol which transiently appears in the cell membrane, as a consequence of receptor-mediated and voltage-dependent hydrolysis of inositol phospholipids. The link of the phosphoinositide system to PKC seems to be that of a major second messenger system in the central nervous system and the possible involvement in significant steps in synaptic transmission have to be given attention. Until recently, PKC was thought to be a single entity, however, molecular cloning analysis has shown that there is a family of PKC-related genes(5–11). At least four subspecies of PKC (α, βI, βII and γ) are expresssed in the mammalian brain. Enzymatically, three major isozymes of PKC (types I, II and III) have been resolved from rat brains, using hydroxyapatite column chromatography(12). All subtypes were activated by diacylglycerol as well as tumor-promoting phorbol esters in the presence of Ca2+ and phosphatidylserine. They bound radioactive phorbol ester and showed similar kinetic properties, although the enzyme having the β-sequence was active at very low concentrations of Ca (8,13). The amino acid sequence of these three types has been identified by comparison with the enzyme separately expressed in the COS cells transfected by respective cDNAs-containing plasmids (14). Type I corresponds to the enzyme encoded with the γ-sequence. Type II is a mixture of βI- and βII-subspecies derived from alternative splicing of a single gene. Type III corresponds to α-subspecies. The αI-and βII-subspecies differ by only about 50 amino acid residues in the carboxyl-terminal regions, show similar kinetic properties and are not separable by conventional enzyme purification procedures. To characterize the tissue specificity of these subspecies, a specific antibody against each subspecies should be a pertinent tool. We present here the cell specific expression of each subspecies of PKC in the rat brain, determined using subspecies specific antibodies.