The generation of REM sleep and the occurrence of REM sleep-related changes in the control of breathing depend on many neurotransmitters and peptides (acetylcholine, serotonin, norepinephrine, glutamate, GABA, glycine, orexin) whose effects are mediated by multiple neurotransmitter receptors. In order to identify these receptors and determine their relative roles, we microinject various neurotransmitter receptor agonists and antagonists into relevant brain regions and observe changes in hypoglossal nerve activity and sleep-wake behavior.
At the motoneuronal level, the REM sleep- related hypotonia of upper airway motoneurons is caused by a combination of state-specific withdrawal of excitatory effects (disfacilitation), state-specific activation of inhibitory mechanisms, and state-specific modulation of other inputs to motoneurons that by themselves are weakly state-dependent (e.g., respiratory inputs).
The release of selected aminergic neurotransmitters (Norepinephrine [NE] and serotonin [5-HT]) and selected peptides (substance P, TRH) is reduced during sleep, and REM sleep in particular. Concurrently, there is an increased release of inhibitory amino acids, such as glycine and GABA, although the contribution of active inhibition to REM sleep-related decrements of upper airway motor tone is small (Kubin, Compr. Physiol. 6:1801-1850, 2016; see: http://dx.doi.org/10.1002/cphy.c160002).
The hypoglossal motor nucleus contains multiple receptors for 5-HT and NE (adrenergic receptors). Because withdrawal of aminergic excitation plays such an important role in the REM sleep-related upper airway hypotonia, it has been important to determine which aminergic receptors mediate these effects. If a given receptor is present in a cell, the cell should express its mRNA. Thus, we have extensively used the technique of single-cell reverse transcription-polymerase chain reaction (RT-PCR) to determine which mRNAs for the at least 11 known 5-HT receptor subtypes and 9 known NE receptor subtypes are present in identified XII motoneurons. We found that mRNAs for two excitatory aminergic receptors, adrenergic alpha1B and serotoneric 5-HT2A, are present in most hypoglossal (XII) motoneurons of adult rats. The inhibitory 5-HT1B receptor is also present in many XII motoneurons, but its role and significance are not yet clear.
We also have extensively used receptor immunohistochemistry to determine which neurotransmitter receptor proteins are expressed in the XII nucleus. 5-HT2A immunostaining is very prominent in the XII nucleus and, indeed, in all orofacial motor nuclei (see PDF).
Notably, the 5-HT2A receptor mRNA and protein levels vary in the XII nucleus with circadian time, such that their level is high at the beginning of the wakefulness/active period and significantly reduced at the beginning of the sleep/rest period (Volgin et al. Respir. Physiol. Neurobiol. 188:301-307, 2013; see: Link). This pattern helps align the availability of these important receptors that drive the use of the tongue with the rest-activity cycle.
Noradrenergic innervation of XII motoneurons originates in a subset of the known groups of pontomedullary noradrenergic neurons (Rukhadze and Kubin, J. Chem. Neuroanat. 33:23-33, 2007; full text: PDF).
One goal of our current studies is to determine whether a selective optogenetic activation of some of these different groups of noradrenergic premotor neurons can enhance motor output to the tongue without concurrent activation of the circuits responsible for termination of sleep (arousal).