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 I-Ca 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 G alpha(0) 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.