Development of G protein-mediated Ca²⁺ channel regulation in mouse embryonic stem cell-derived neurons

Besides other mechanisms, the influx of Ca²⁺ into embryonic neurons controls growth and differentiation processes. To study the expression and regulation of voltage-gated Ca²⁺ channels during early neurogenesis, we measured whole-cell Ca²⁺ currents (I_Ca) in neurons developing from pluripotent embryonic stem cells. Various receptor agonists, including somatostatin and baclofen, reversibly inhibited lca in embryonic stem cell-derived neurons. The effects of somatostatin and baclofen were abolished by pretreatment of cells with pertussis toxin and mimicked by intracellular infusion of guanosine 5′-O-(3-thiotriphosphate), suggesting the involvement of pertussis toxin-sensitive G proteins in I_Ca inhibition. Investigations at different stages of neuronal differentiation showed that somatostatin efficiently suppressed L- and N-type Ca²⁺ channels in immature as well as mature neurons. In contrast, inhibition of L- and N-type channels by baclofen was rarely observed at the early stage. In terminally differentiated neurons, responses to baclofen were as prominent as those to somatostatin but were confined to N-type Ca²⁺ channels. The stage-dependent sensitivity of voltage-gated Ca²⁺ channels to somatostatin and baclofen was not due to differential expression of Goto isoforms, as revealed by reverse transcription-polymerase chain reaction and immunofluorescence microscopy. These findings demonstrate that specific neurotransmitters such as somatostatin regulate voltage-gated Ca²⁺ channels via G proteins during the early stages of neurogenesis, thus providing a mechanism for the epigenetic control of neuronal differentiation.