DEVELOPMENT OF G-PROTEIN-MEDIATED CA2-DERIVED NEURONS( CHANNEL REGULATION IN MOUSE EMBRYONIC STEM CELL)

Besides other mechanisms, the influx of Ca2+ into embryonic neurons co ntrols growth and differentiation processes. To study the expression a nd regulation of voltage-gated Ca2+ channels during early neurogenesis , we measured whole-cell Ca2+ currents (I-Ca) in neurons developing fr om pluripotent embryonic stem cells, Various receptor agonists, includ ing somatostatin and baclofen, reversibly inhibited I-Ca in embryonic stem cell-derived neurons. The effects of somatostatin and baclofen we re abolished by pretreatment of cells with pertussis toxin and mimicke d by intracellular infusion of guanosine 5'-O-(3-thiotriphosphate), su ggesting the involvement of pertussis toxin-sensitive G proteins in I- Ca inhibition. Investigations at different stages of neuronal differen tiation showed that somatostatin efficiently suppressed L- and N-type Ca2+ channels in immature as well as mature neurons. In contrast, inhi bition of L- and N-type channels by baclofen was rarely observed at th e early stage, In terminally differentiated neurons, responses to bacl ofen 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 transcripti on-polymerase chain reaction and immunofluorescence microscopy. These findings demonstrate that specific neurotransmitters such as somatosta tin regulate voltage-gated Ca2+ channels via G proteins during the ear ly stages of neurogenesis, thus providing a mechanism for the epigenet ic control of neuronal differentiation.