Synaptogenesis of cultured striatal neurons in serum-free medium: A morphological and biochemical study (neuronal differentiation/synapse formation/synapsin I/signal transduction/y-aminobutyric acid release)

Striatal neurons were cultured from the fetal mouse brain and maintained in serum-free medium for 14-21 days in vitro (DIV). Pretreatment of the culture dishes succes- sively with a polycation followed by fetal calf serum resulted in rapid neuron attachment and neurite proliferation. After 9-10 DIV, electron microscope observations revealed the presence of vesicles in axon terminals forming mature synapses with axons and perikarya of adjacent neurons and in varicosities along extended axons. Synapsin I, a synaptic vesicle-specific protein, was present only in neuronal perikarya after 3 DIV, in perikarya and in varicosites along extended axons after 6 DIV, and in varicosities and contact points between axon terminals and adjacent axons or perikarya after 11-14 DIV. Neurotrans- mitter-stimulated intracellular formation of cAMP decreased markedly during neuronal differentiation. Inositol phosphate formation in response to neurotransmitters, however, in- creased significantly throughout the period of striatal neuronal development. K+ (56 mM) depolarization resulted in a 2-fold increase in endogenous y-aminobutyric acid (GABA) release from striatal neurons, 50% of which was Ca2+-dependent, between 3 and 11 DIV. Between 11 and 14 DIV, subsequent to synapse formation (as revealed by electron microscope obser- vations), GABA release evoked by 56 mM K+ increased up to 5-fold, 75 % of which was Ca2+-dependent. It appears that the complete differentiation of striatal neurons in serum-free medium may provide a suitable model for the study of the physiological and regulatory mechanisms involved in nerve cell development.