SYNAPTOGENESIS AND DISTRIBUTION OF PRESYNAPTIC AXONAL VARICOSITIES INLOW-DENSITY PRIMARY CULTURES OF NEOCORTEX - AN IMMUNOCYTOCHEMICAL STUDY UTILIZING SYNAPTIC VESICLE-SPECIFIC ANTIBODIES, AND AN ELECTROPHYSIOLOGICAL EXAMINATION UTILIZING WHOLE-CELL RECORDING

Low-density primary cultures of neocortical neurons were utilized to e xamine: (i) early interactions of growing neurites with morphological characteristics of axons with other neuronal elements, and (ii) the di stribution of presynaptic axonal varicosities closely apposed to MAP-2 immunoreactive, putatively postsynaptic, dendrites. At the Light micr oscopical level axonal varicosites, presumably presynaptic terminals, were identified using immunocytochemistry incorporating antibodies spe cific for the synaptic vesicle antigens synaptophysin and synapsin. Th e presence of synaptophysin- and synapsin-immunoreactive swellings alo ng axonal processes was first detected at 5 days post-plating and was also apparent in axons growing in isolation. At 5-7 days in vitro, imm unolabelled axonal varicosities in close apposition to putative postsy naptic dendrites (MAP-2 immunoreactive) dendrites were detected. Elect rophysiologically active synaptic contacts can also readily be detecte d at this stage. After 3 weeks in vitro presynaptic contacts do appear to be distributed heterogeneously along postsynaptic dendrites of man y neurons in culture. As the culture matures a higher number of presyn aptic profiles can be seen along dendrites, with a centrifugal distrib ution, e.g. a higher density of presynaptic axonal terminals in close apposition to more distal regions of larger dendrites, putatively cons idered to be apical dendrites of pyramidal-like neurons. In our cultur es, the overall increase in the density and the pattern of distributio n of presynaptic axon terminals immunoreactive for synaptic vesicle an tigens closely apposed to putative post-synaptic structures mimics the general postnatal increase of synaptic density in the neocortex in vi vo. Thus, low density primary cultures of neocortical neurons offer a valuable system to explore and manipulate (i) the molecular and cellul ar basis of neocortical synaptogenesis, and (ii) the pharmacology of n eocortical synaptic transmission.