LAUSR

The sleep-related withdrawal of noradrenergic excitation and increased cholinergic inhibition of hypoglossal motoneurons (HM), which control the tone of the GG, the largest protruder muscle of the tongue, play a critical role in obstructive sleep apnea pathophysiology. In an animal model of REM sleep, antagonism of alpha1 noradrenergic signaling by microinjections of prazosin into the hypoglossal nucleus (HN) strongly decreased hypoglossal nerve activity. However, a significant delay of this decrease prompted a hypothesis that prazosin must diffuse outside the HN to block alpha1-adrenoceptors that are located on interneurons, which mediate noradrenergic drive to HM. Here, we aimed to map the PHR by microinjecting alpha1 drugs for possible location of these interneurons. Adult C57bl/6J mice (n=28) of both sexes were anesthetized with either ketamine/xylazine (120/20 mg/kg, i.p.) or 1.5-2.0% isoflurane inhaled through a nose mask. Electromyogram electrodes were implanted into GG and diaphragm to record their activity. We microinjected phenylephrine (alpha1-adrenoceptor agonist, 5-10 nl, 1 mM) in both ketamine/xylazine- and isoflurane-anesthetized mice, while prazosin (10 nl, 0.2 mM) was injected only in mice under isoflurane anesthesia. The injection sites were marked with Pontamine Skye Blue. After the completion of experiments, all animals were perfused, and brains removed for histological analysis. The activity of GG showed spontaneous phasic inspiratory modulation with minimal tonic component under both anesthesia conditions although its amplitude was markedly lager in animals anesthetized by isoflurane. Effective microinjections of phenylephrine elicited short-lasting (approximately 15 minutes) excitatory GG responses. When injected into the HN, phenylephrine strongly excited GG as compared to injections to the PHR in ketamine/xylazine-anesthetized mice. However, in isoflurane-anesthetized animals, microinjections of phenylephrine into the PHR induced greater activation of GG than injections onto the HN. Microinjections of prazosin in isoflurane-anesthetized mice produced long-lasting disfacilitation of GG activity. However, prazosin injections into the PHR elicited responses with the shortest latencies (0-120 s). These results suggest that the putative interneurons that mediate noradrenergic drive to HM are located in the PHR. This finding is important for deciphering the neuronal network that controls the state-dependent activity of GG, which may provide new therapeutic targets for OSA treatment. NIHR01HL133847, NIAR01AG065233.