Multiple voltage-sensitive calcium channels are probably involved in endogenous GABA release from striatal neurones differentiated in primary culture

Abstract

Calcium-dependent release of neurotransmitters is thought to be due to Ca²⁺ entry into nerve terminals, but the identities of the various voltage-sensitive Ca²⁺ channels (VSCC) involved in this process remain obscure. To elucidate the types of VSCCs involved in the release process, we studied the effects of various organic Ca²⁺ channel antagonists and agonists on the release of endogenous γ-aminobutyric acid (GABA) from mouse striatal neurones differentiated in primary culture. Diltiazem, verapamil and methoxyverapamil (D 600) inhibited K⁺-evoked (30 mM) GABA release at very high concentrations (> 1 μM). The dihydropyridine (DHP) nifedipine, at low concentrations (0.01–1.00 μM), was able to inhibit part of the K⁺-evoked GABA release (25.6±7.3% inhibition at 1 μM). This is in agreement with the high affinity of nifedipine for DHP binding sites. The DHPs, BAY K 8644 (EC₅₀ = 41±15 nM) and CGP 28.392, which possess agonist properties at VSCCs, increased the 15 mM K⁺-evoked GABA release. The release evoked by the combination of K⁺ (15 μM) and BAY K 8644 (up to 10 μM) remained smaller than the release elicited by 30 mM K⁺. The effect of BAY K 8644 (1 μM) was inhibited by nifedipine (IC₅₀ 0.55±0.05 μM). When Na⁺ ions were replaced by choline, basal and K⁺-evoked GABA release was significantly increased. Even in the absence of external Na+, nifedipine (1 μM) was not able to totally block the K⁺ effect. Moreover amiloride, a drug known to inhibit Na⁺/Ca⁺ exchange, and tetrodotoxin (TTX), did not modify the 30 mM K⁺ response. Therefore, nifedipine-insensitive K⁺-evoked GABA release is not due to Na⁺-dependent Ca²⁺ entry. These results can be explained by the presence of DHP-sensitive and insensitive Ca²⁺ channels on nerve terminals, each involved in the release process.