Muscarinic M1 modulation of N and L types of calcium channels is mediated by protein kinase C in neostriatal neurons

In some neurons, muscarinic M1-class receptors control L-type (CaV1) Ca2+-channels via protein kinase C (PKC) or calcineurin (phosphatase 2B; PP-2B) signaling pathways. Both PKC and PP-2B pathways start with phospholipase C (PLC) activation. In contrast, P/Q- and N-type (CaV2.1, 2.2, respectively) Ca2+-channels are controlled by M2-class receptors via G proteins that may act, directly, to modulate these channels. The hypothesis of this work is that this description is not enough to explain muscarinic modulation of Ca2+ channels in rat neostriatal projection neurons. Thus, we took advantage of the specific muscarinic toxin 3 (MT-3) to block M4-type receptors in neostriatal neurons, and leave in isolation the M1-type receptors to study them separately. We then asked what Ca2+ channels are modulated by M1-type receptors only. We found that M1-receptors do modulate L, N and P/Q-types Ca2+ channels. This modulation is blocked by the M1-class receptor antagonist (muscarinic toxin 7, MT-7) and is voltage-independent. Thereafter, we asked what signaling pathways, activated by M1-receptors would control these channels. We found that inactivation of PLC abolishes the modulation of all three channel types. PKC activators (phorbol esters) mimic muscarinic actions, whereas reduction of intracellular calcium virtually abolishes all modulation. As expected, PKC inhibitors prevented the muscarinic reduction of the afterhyperpolarizing potential (AHP), an event known to be dependent on Ca2+ entry via N- and P/Q-type Ca2+ channels. However, PKC inhibitors (bisindolylmaleimide I and PKC-1936) only block modulation of currents through N and L types Ca2+ channels; while the modulation of P/Q-type Ca2+ channels remains unaffected. These results show that different branches of the same signaling cascade can be used to modulate different Ca2+ channels. Finally, we found no evidence of calcineurin modulating these Ca2+ channels during M1-receptor activation, although, in the same cells, we demonstrate functional PP-2B by activating dopaminergic D2-receptor modulation.