Involvement of the Voltage-Gated Calcium Channels L- P/Q- and N-Types in Synapse Elimination During Neuromuscular Junction Development

Abstract

During the nervous system development, synapses are initially overproduced. In the neuromuscular junction (NMJ) however, competition between several motor nerve terminals and the synapses they made ends with the maturation of only one axon. The competitive signaling between axons is mediated by the differential activity-dependent release of the neurotransmitter ACh, co-transmitters, and neurotrophic factors. A multiple metabotropic receptor-driven downstream balance between PKA and PKC isoforms modulates the phosphorylation of targets involved in transmitter release and nerve terminal stability. Previously, we observed in the weakest endings on the polyinnervated NMJ that M1 mAChR receptors reduce ACh release through the PKC pathway coupled to an excess of Ca2+ inflow through P/Q- N- and L-type voltage-gated calcium channels (VGCC). This signaling would contribute to the elimination of this nerve terminal. Here, we investigate the involvement of the P/Q-, N-, and L-subtype channels in transgenic B6.Cg-Tg (Thy1-YFP)16-Jrs/J mice during synapse elimination. Then, the axon number and postsynaptic receptor cluster morphologic maturation were evaluated. The results show that both L- and P/Q-type VGCC (but not the N-type) are equally involved in synapse elimination. Their normal function favors supernumerary axonal loss by jointly enhancing intracellular calcium [Ca2+]i. The block of these VGCCs or [Ca2+]i i sequestration results in the same delay of axonal loss as the cPKCβI and nPKCε isoform block or PKA activation. The specific block of the muscle cell’s contraction with μ-conotoxin GIIIB also delays synapse maturation, and thus, a retrograde influence from the postsynaptic site regulating the presynaptic CaV1.3 may contribute to the synapse elimination.

Regulation of pain neurotransmitters and chondrocytes metabolism mediated by voltage-gated ion channels: A narrative review

“…They are divided into six subtypes, L, T, N, P, Q, and R, based on the conductivity of calcium channels and the voltage sensitivity [ 23 ]. VSCCs are located on the cell membrane of all excitable cells, such as neurons, muscle cells, and gland cells, and are mainly responsible for triggering the release of transmitters and regulating cell metabolism [ 24 ]. The action potential of neurons depends on N, P, Q, and R types.…”

Muscarinic Receptors in Developmental Axonal Competition at the Neuromuscular Junction

“…It is known that reduction of the postsynaptic activity or contraction results in a delay in synapse elimination during NMJ development [ 18 , 92 – 95 ]. In line with this, we made experiments blocking the muscle cell’s contractile activity with μ-conotoxin GIIIB which blocks muscle cell sodium channel but preserves neurotransmission because does not influence the nAChR [ 26 ]. Accordingly, incubation with μ-conotoxin GIIIB also results in a delay in axon loss.…”

“…The protein expression of all these channels increases during the development P5-P7 time period. Western blots at P30 show that the P/Q level is at its highest whereas α1D-L and N channel proteins stabilize at a lower level [ 26 ].…”

“…The L and P/Q-type (but not the N-type) channels tonically enhance synapse elimination because their block prolongs the multiinnervation of the developing NMJ, whereas their exogenous stimulation (Bay-K8644 for L channel and GV-58 for P/Q channels) results in a significant acceleration [ 26 ]. Moreover, the delaying effect on axon loss of the L and P/Q channels block is equal to that produced by intracellular calcium sequestration with BAPTA-AM.…”

“…During development, skeletal myocytes start polyinnervated by several axons [15][16][17], but after their competitive interactions, neuromuscular junctions (NMJ) finally become innervated by only one axon [6,13,14,18]. There are many reviews about synapse elimination mainly focused on the NMJ [1,[18][19][20][21][22][23][24][25][26][27]. Several relevant cues of the molecular and cellular mechanisms involved in the elimination of supernumerary nerve terminals have been investigated and collected in the cited reviews.…”