Photocontrol of muscarinic receptor activity in the mouse somatosensory cortex monitored by two-photon calcium imaging

Muscarinic acetylcholine receptors (mAChRs) contribute to both the facilitation and inhibition of cortical activity, with a role in attention, wakefulness and sleep states regulation. Acetylcholine is sensed by extrasynaptic mAChRs, which are present in all cortical layers where they modulate the excitability of pyramidal cells. Current pharmacological tools that regulate neural activity lack tissue selectivity and affect the entire nervous system and thus cannot be used to activate or inhibit selective regions on demand. On the other hand, photostimulation with optogenetics requires gene manipulation, which poses hurdles for clinical translation. Photopharmacology, the development of molecules with photoswitchable moieties that can be activated and deactivated with illumination, allows focalized switching, reduce adverse effects and enhances efficacy. Here we characterise a photoswitchable muscarinic activator derivative that is inactive in the dark and activates mAChRs when they are illuminated with tissue-penetrating amber light (590 nm). We combined the use of this light-responsive effector with two-photon calcium imaging to optically control neuron activity of the somatosensory cortex layer II/III. We demonstrate that a photoswitchable molecule and the application of patterns of light can be used to control drug activity on demand. Muscarinic activation with 590 nm light produced a clear difference in activity, whereas no changes in calcium activity were observed in the control group for the same illumination cycles. All mice recovered without behaviour impairments, which demonstrates that this compound does not produce acute toxicity. This photoswitchable drug is thus a unique tool to optically control muscarinic endogenous receptors in brain tissue in vivo and without genetic manipulation.