Afferent-specific modulation of excitatory synaptic transmission by acetylcholine and serotonin in the prelimbic cortex

Acetylcholine (ACh) and serotonin (5-hydroxytryptamine, or 5-HT) differentially regulate the excitability of pyramidal neurons in the mouse prelimbic (PL) cortex according to their long-distance projections. Here we tested for afferent- and/or target-specific modulation of glutamate release by ACh and 5-HT in two long-distance excitatory projections to the PL cortex: commissural (COM) afferents from the contralateral cortex and projections from the mediodorsal nucleus (MDN) of the thalamus. Using ex-vivo optogenetic approaches, we mapped the connectivity and neuromodulation of COM and MDN afferents in layer 5 intratelencephalic (IT) and extratelencephalic (ET) projection neurons. Dual whole-cell recordings in pairs of IT and ET neurons revealed that COM afferents target both neuron subtypes, but that MDN afferents selectively target IT neurons. Both afferents exhibited similar target-independent short-term synaptic plasticity (paired-pulse facilitation) across a range of frequencies, but were differentially modulated by ACh and 5-HT. In both control conditions and after isolating monosynaptic connections with tetrodotoxin and 4-aminopyridine, COM transmission was suppressed strongly by ACh and moderately by 5-HT, while MDN transmission was largely unaffected by either neuromodulator. Suppression o,f COM transmission by ACh or 5-HT was concentration dependent and mediated by M4 muscarinic or 5-HT1B receptors, respectively. Chemogenetic inhibition of hM4Di-expressing COM terminals mimicked the suppressive effects of ACh and 5-HT on synaptic transmission. Our results demonstrate that ACh and 5-HT preferentially regulate COM synaptic transmission, albeit to different degrees, and suggest that, through their combined pre- and postsynaptic neuromodulation, ACh and 5-HT may differentially regulate cortico-striatal-thalamic loops to influence cognition and behavior.