Synaptic activation of Ca2+ action potentials in immature rat cerebellar granule cells in situ.
Although numerous Ca2+ channels have been identified in cerebellar granule cells, their role in regulating excitability remained unclear. We therefore investigated the excitable response in granule cells using whole cell patch-clamp recordings in acute rat cerebellar slices throughout the time of development (P4-P21, n = 183), with the aim of identifying the role of Ca2+ channels and their activation mechanism. After depolarizing current injection, 46% of granule cells showed Ca2+ action potentials, whereas repetitive Na+ spikes were observed in an increasing proportion of granule cells from P4 to P21. Because Ca2+ action potentials were no longer observed after P21, they characterized an immature granule cell functional stage. Ca2+ action potentials consisted of an intermediate-threshold spike (ITS) activating at -60/-50 mV and sensitive to voltage inactivation and of a high-threshold spike (HTS), activating at above -30 mV and resistant to voltage inactivation. Both ITS and HTS comprised transient and protracted Ca2+ channel-dependent depolarizations. The Ca2+ action potentials could be activated synaptically by excitatory postsynaptic potentials, which were significantly slower and had a proportionately greater N-methyl-D-aspartate (NMDA) receptor-mediated component than those recorded in cells with fast repetitive Na+ spikes. The NMDA receptor current, by providing a sustained and regenerative current injection, was critical for activating the ITS, which was not self-regenerative. Moreover, NMDA receptors determined temporal summation of impulses during repetitive mossy fiber transmission, raising membrane potential into the range required for generating protracted Ca2+ channel-dependent depolarizations. The nature of Ca2+ action potentials was considered further using selective ion channel blockers. N-, L-, and P-type Ca2+ channels generated protracted depolarizations, whereas the ITS and HTS transient phase was generated by putative R-type channels (R(ITS) and R(HTS), respectively). R(HTS) channels had a higher activation threshold and were more resistant to voltage inactivation than R(ITS) channels. At a mature stage, most of the Ca2+-dependent effects depended on the N-type current, which promoted spike repolarization and regulated the Na+-dependent discharge frequency. These observations relate Ca2+ channel types with specific neuronal excitable properties and developmental states in situ. Synaptic NMDA receptor-dependent activation of Ca2+ action potentials provides a sophisticated mechanism for Ca2+ signaling, which might be involved in granule cell development and plasticity.
D'Angelo
E
E
Instituto di Fisiologia Generale, Pavia, Italy.
De Filippi
G
G
Rossi
P
P
Taglietti
V
V
D000200:Action Potentials / Q000187:drug effects / Q000502:physiology; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D004594:Electrophysiology; D019706:Excitatory Postsynaptic Potentials / Q000187:drug effects / Q000502:physiology; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009412:Nerve Fibers / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D051381:Rats; D017208:Rats, Wistar; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors / Q000502:physiology; D013569:Synapses / Q000187:drug effects / Q000502:physiology*
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D016194:Receptors, N-Methyl-D-Aspartate