Muscarinic modulation of synaptic transmission via endocannabinoid signalling in the rat midbrain periaqueductal grey

The midbrain periaqueductal grey (PAG) is involved in organizing behavioural responses to threat, stress and pain. These PAG functions are modulated by cholinergic agents. In the present study, we examined the cholinergic modulation of synaptic transmission in the PAG using whole-cell voltage-clamp recordings from rat midbrain slices. We found that the cholinergic agonist carbachol reduced the amplitude of evoked inhibitory and excitatory postsynaptic currents (IPSCs, EPSCs) in all PAG neurons and this was abolished by the muscarinic receptor antagonist atropine. Carbachol increased the paired-pulse ratio of evoked IPSCs and EPSCs, and reduced the rate, but not the amplitude of spontaneous miniature IPSCs. The carbachol inhibition of evoked IPSCs was mimicked by the acetylcholinesterase inhibitor physostigmine and was reduced by the M1 and M1/M3 muscarinic cholinergic receptor (mAChR) antagonists pirenzepine and 4DAMP, but not by the M2 and M4 antagonists gallamine and PD-102807. The carbachol inhibition of evoked IPSCs was reduced by the cannabinoid CB1 receptor antagonist AM251 and the DAG lipase inhibitor tetrahydrolipstatin, and was abolished in the presence of both AM251 and gallamine. The carbachol inhibition of evoked EPSCs was also reduced in the presence of both gallamine and AM251. These results indicate that M1 induced inhibition of GABAergic transmission within the PAG is mediated via endocannabinoids which are produced via the phospholipase C/DAG lipase pathway and activate presynaptic cannabinoid CB1 receptors. Thus, presynaptic muscarinic modulation of PAG function is mediated indirectly by M1 receptor induced endocannabinoid-signalling and directly by M2 receptors.