Perinatal mAChR-mediated activation of cortical subplate neurons elicits network activity driving basket cell...

The activation of mAChR on subplate neurons has been shown to trigger oscillatory glutamatergic network activity in early postnatal neocortex. Here, we explored the long-term consequences of subplate activation on the differentiation of cortical neurons. Stimulating newborn somatosensory and visual cortex ex vivo and organotypic cortex cultures (OTCs) of the visual cortex with carbachol (CCH) led to an upregulation of Bdnf and Zif-268. In OTCs, the stimulation increased the amplitude and frequency of calcium events in the gray matter layers at DIV 3-5. At DIV 5, an acute stimulation rapidly activates p21ras; increases ERK phosphorylation; induces Bdnf, c-Fos, and Zif-268 expression; and increases GluN1, GluN2A, GluN2B, PSD-95, GAD-65, and synaptophysin protein expression. The NMDA receptors were functional, as shown by enhanced ligand-induced dendritic injury. Repeated DIV 1-5 mAChR activation increased, at DIV 10, the expression of GluN2A, of an essential protein of the inhibitory presynapse, VGAT, and the basket-cell-specific presynaptic calcium sensor synaptotagmin-2. Furthermore, it increased the frequency of interneuronal calcium events at DIV 10-15, dendritic length of basket cells, and accelerated the local arborization of basket cell axons at DIV 15. The results suggest that temporally limited mAChR-mediated activation of subplate neurons evokes glutamatergic network activity, which, gradually over the following weeks, promotes cortical circuit development. Specifically, it enhances the expression of excitatory and inhibitory synaptic proteins and accelerates the morphological and functional maturation of basket cells, highlighting the cholinergic input to subplate neurons as a critical regulator of interneuron development.