Pharmacological characterisation of voltage-sensitive calcium channels and neurotransmitter release from mouse cerebellar granule cells in culture

Using subtype-specific Ca-channel blockers, we have characterised the voltage-sensitive Ca²⁺ currents as well as neurotransmitter release from cultured mouse cerebellar granule cells. The whole cell version of the patch clamp technique was adapted to monitor the isolated Ca-channel currents. The currents were activated at potentials more positive than −40 mV and were composed of at least four pharmacological distinct components being sensitive to nifedipine (35%), ω-conotoxin GVIA (10%), and ω-agatoxin IVA (42%) corresponding to L-, N-, and P-channel-mediated currents. The insensitive fraction (13%) possibly represented R channels. High potassium-evoked release of ³H-D-aspartate was used as a model of synaptic release. These studies were performed at relatively mild stimulation conditions (30 mM K⁺, 0.4 mM Ca²⁺), and 85% of the evoked release was Ca²⁺ dependent as well as tetrodotoxin and Cd²⁺ sensitive. Nifedipine and ω-agatoxin IVA dose dependently (IC₅₀ values of 10 nM and 0.7 nM, respectively) blocked most of the release, whereas ω-conotoxin MVIIA (IC₅₀ = 5 nM) caused partial blockage. The results indicate that several subtypes of voltage-sensitive Ca channels are present in mouse cerebellar granule cells. Furthermore, the data suggest that L, N, and P channels act in concert in the neurotransmitter release process. J. Neurosci. Res. 48:43–52, 1997. © 1997 Wiley-Liss, Inc.