DREADD-induced activation of subfornical organ neurons stimulates thirst and salt appetite

The subfornical organ (SFO) plays a pivotal role in body fluid homeostasis through its ability to integrate neurohumoral signals and subsequently alter behavior, neuroendocrine function, and autonomic outflow. The purpose of the present study was to evaluate whether selective activation of SFO neurons using virally mediated expression of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) stimulated thirst and salt appetite. Male C57BL/6 mice (12-15 wk) received an injection of rAAV2-CaMKII-HAhM3D( Gq)-IRES-mCitrine targeted at the SFO. Two weeks later, acute injection of clozapine N-oxide (CNO) produced dose-dependent increases in water intake of mice with DREADD expression in the SFO. CNO also stimulated the ingestion of 0.3 M NaCl. Acute injection of CNO significantly increased the number of Fos-positive nuclei in the SFO of mice with robust DREADD expression. Furthermore, in vivo single-unit recordings demonstrate that CNO significantly increases the discharge frequency of both ANG II- and NaCl-responsive neurons. In vitro current-clamp recordings confirm that bath application of CNO produces a significant membrane depolarization and increase in action potential frequency. In a final set of experiments, chronic administration of CNO approximately doubled 24-h water intake without an effect on salt appetite. These findings demonstrate that DREADD-induced activation of SFO neurons stimulates thirst and that DREADDs are a useful tool to acutely or chronically manipulate neuronal circuits influencing body fluid homeostasis.