REVERSIBLE PHOTOACTIVATION OF ENDOGENOUS NICOTINIC RECEPTORS IN REWARD AND PAIN CIRCUITS OF BEHAVING MICE

Nicotinic acetylcholine receptors (nAChRs) are crucial modulators of neural circuits and play key roles in reward, motivation, and pain modulation. However, dissecting how nAChR activation controls signaling in defined circuits in vivo remains challenging due to widespread cholinergic innervation and spatiotemporal imprecision of systemic pharmacology.

We developed high-affinity, photoactivatable nAChR agonists that enable temporally and spatially precise manipulation of nicotinic signaling in the intact brain. Using functional ultrasound imaging, we show that these caged compounds can be systemically administered in their inactive form via a single intraperitoneal injection, and repeatedly photoactivated in targeted regions via implanted optical fibers.

Optical uncaging in the ventral tegmental area (VTA) produced neuronal activation and rapid increases in nucleus accumbens dopamine, demonstrating circuit-specific control of mesolimbic output. A single systemic injection enabled repeated, reliable stimulation for up to five hours, allowing behavioral assessment in freely moving mice without physical intervention. Uncaging in the VTA, but not in adjacent regions, elicited a robust, real-time preference for the stimulation-paired compartment.

We further investigated nAChR roles in pain modulation by optically uncaging nicotinic agonists in the periaqueductal gray (PAG), a key region for descending analgesic control. PAG uncaging produced behavioral changes in nociceptive responses, demonstrating that spatially restricted nAChR activation modulates pain processing.

Together, these results establish systemic photopharmacology of nAChRs as a method for precise, repeatable in vivo control of cholinergic signaling. Our approach provides a powerful tool to dissect nicotinic receptor function across behavioral domains and is extensible to other receptor systems and therapeutics.