Ultrastructural evidence for synaptic interactions between thalamocortical axons and subplate neurons

Thalamic axons are known to accumulate in the subplate for a protracted period prior to invading the cortical plate and contacting their ultimate targets, the neurons of layer 4. We have examined the synaptic contacts made by visual and somatosensory thalamic axons during the transition period in which axons begin to leave the subplate and invade the cortical plate in the ferret. We first determined when geniculocortical axons leave the subplate and begin to grow into layer 4 of the visual cortex by injecting 1,1'-dioctadecyl-3,3,3',3'-tetramethyl indocarbocyanine (Dil) into the lateral geniculate nucleus (LGN). By birth most LGN axons are still confined to the subplate. Over the next 10 days LGN axons grow into layer 4, but many axons retain axonal branches within the subplate. To establish whether thalamic axons make synaptic contacts within the subplate, the anterograde tracer PHA-L was injected into thalamic nuclei of neonatal ferrets between postnatal day 3 and 12 to label thalamic axons at the electron microscope level. The analysis of the PHA-L injections confirmed the Dil data regarding the timing of ingrowth of thalamic axons into the cortical plate. At the electron microscope level, PHA-L-labelled axons were found to form synaptic contacts in the subplate. The thalamic axon terminals were presynaptic primarily to dendritic shafts and dendritic spines. Between postnatal days 12 and 20 labelled synapses were also observed within layer 4 of the cortex. The ultrastructural appearance of the synapses did not differ significantly in the subplate and cortical plate, with regard to type of postsynaptic profiles, length of postsynaptic density or presynaptic terminal size. These observations provide direct evidence that thalamocortical axons make synaptic contacts with subplate neurons, the only cell type within the subplate possessing mature dendrites and dendritic spines; they also suggest that functional interactions between thalamic axons and subplate neurons could play a role in the establishment of appropriate thalamocortical connections.