DEVELOPMENT OF THE GABAERGIC PHENOTYPE IN MURINE SPINAL-CORD DORSAL-ROOT GANGLION CULTURES

Murine spinal cord and dorsal root ganglion GABAergic neurons, derived from 12-day-old fetuses, were examined autoradiographically, biochemi cally and immunocytochemically in vitro to determine the timecourse of appearance and maturation of this phenotype and the extent and mode o f its innervation of target neurons. Specific H-3-GABA uptake into spi nal cord neurons was the first property to develop and was present at the earliest time studied, one day in vitro. Immunocytochemical locali zation of glutamic acid decarboxylase (GAD) revealed positively staine d neurons beginning at four days. At five days in vitro, electron micr oscopic immunocytochemistry revealed GAD-immunoreactive (GAD-IMR) bout ons investing neuronal perikarya as well as neuronal processes. By one week in vitro, GAD-IMR neurons constituted 27% of the total populatio n and GAD-IMR boutons could be seen contacting every cell with a neuro nal morphology. The mode of investment of target neurons by GAD-IMR bo utons was not circumscribed to either soma or dendrites but usually in volved the entire neuronal perimeter and did not change with time in c ulture. Three morphologically distinct types of GAD-IMR neurons were e vident: a small, bipolar type; a medium-sized multipolar neuron which was the most common and a large, multipolar type, resembling a motoneu ron. A small population (8%) of dorsal root ganglion neurons was found to contain GAD both biochemically and immunocytochemically but was ne ver invested by GAD-IMR boutons. GAD activity in vitro paralleled in v ivo levels with maximal activity being reached at four weeks in vitro and 10 days postnatally in the intact mo use spinal cord. Murine spina l cord GABAergic neurons are a morphologically diverse and abundant ne uronal population with extensive, precocious innervation of all other neuronal phenotypes in vitro suggesting that GABA has a widespread inf luence over other developing neuronal systems in the murine spinal cor d.