Neural Control of Respiration by the Retrotrapezoid Nucleus.

Basting, Tyler, Neuroscience - Graduate School of Arts and Sciences, University of Virginia
Winckler, Bettina, Department of Cell Biology, University of Virginia
Bayliss, Douglas, Department of Pharmacology, University of Virginia
Corwin, Jeffrey, Department of Neuroscience, University of Virginia
Perez-Reyes, Edward, Department of Pharmacology, University of Virginia
Stornetta, Ruth, Department of Pharmacology, University of Virginia
Guyenet, Patrice, Department of Pharmacology, University of Virginia

Rationale and objectives: Central respiratory chemoreceptors (CRCs) detect brain PaCO2 and adjust lung ventilation to maintain PaCO2 and pH constant regardless of the absolute level of lung ventilation. They perform this function in cooperation with the carotid bodies, sensory organs that respond to hypoxia in a pH-dependent manner. The existence of CRCs has been known for over a century but their cellular nature and location have remained highly controversial until recently. The retrotrapezoid nucleus (RTN), a collection of ~2000 glutamatergic neurons in rats that are activated by hypercapnia in vivo and by acidification in slices, had been identified as a CRC candidate just prior to my PhD work. My first objective was to seek additional and critical evidence that RTN neurons are CRCs by determining whether these neurons stimulate breathing in proportion to arterial PCO2 in intact unanesthetized rats. Having verified that this was the case I tested a logical corollary of this theory namely that RTN is silenced under hypoxic conditions due to respiratory alkalosis. Then I proceeded to test whether RTN neurons sustain breathing automaticity during sleep. Finally, I tested whether RTN compensates for the lack peripheral chemoreceptor input.

Methods: RTN neurons were bilaterally transduced to express the proton pump archaerhodopsin using a lentiviral vector. Using anesthetized rats, I confirmed that RTN neurons were silenced by activating archaerhodopsin with green light. I examined the effect of short periods of RTN inhibition (10s) on breathing (plethysmography), EEG and neck EMG in conscious rats in which arterial pH was changed by exposing the animals to various FiO2 levels alone or with the addition of 3% FiCO2 or acetazolamide (ACTZ). Rats were studied in different states of vigilance (quiet awake/non-REM/REM). Arterial blood gases (PaO2, PaCO2), pHa and HCO3 were measured under each condition.

Results: RTN inhibition (bilateral) reduced breathing frequency and amplitude in direct proportion to arterial plasma pH (pHa) below a threshold of 7.53. Above this level, RTN inhibition had no effect suggesting that the nucleus was silent. In normoxia, RTN inhibition reduced breathing equally during non-REM sleep and quiet wake. By contrast RTN inhibition had very little effect on breathing during REM sleep.

Conclusions: RTN drives breathing in direct proportion to arterial PCO2 in intact unaesthetized rats, consistent with the theory that these neurons are CRCs. RTN neurons are silent above pHa 7.5 and increasingly active below this value. RTN regulates breathing automaticity about equally during non-REM sleep and quiet waking, conditions under which the respiratory pattern generator is autorhythmic. By contrast, RTN has a much reduced influence on breathing during REM sleep, consistent with the known reduction of the chemoreflex during this stage of sleep. Hyperoxia activates RTN, which maintains ventilation at control level (normoxia) despite the presumed loss of carotid body input. Finally we demonstrate that RTN and the carotid bodies can regulate breathing independently of each other.

PHD (Doctor of Philosophy)
breathing, retrotrapezoid nucleus
CC0 (permitting unconditional free use, with or without attribution)
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