Combined unilateral blockade of cholinergic, peptidergic, and serotonergic receptors in the ventral respiratory column does not affect breathing in awake or sleeping goats

Abstract

Previous work in intact awake and sleeping goats has found that unilateral blockade of excitatory inputs in the ventral respiratory column (VRC) elicits changes in the concentrations of multiple neurochemicals, including serotonin (5-HT), substance P, glycine, and GABA, while increasing or having no effect on breathing. These findings are consistent with the concept of interdependence between neuromodulators, whereby attenuation of one modulator elicits compensatory changes in other modulators to maintain breathing. Because there is a large degree of redundancy and multiplicity of excitatory inputs to the VRC, we herein tested the hypothesis that combined unilateral blockade of muscarinic acetylcholine (mACh), neurokinin-1 (NK1, the receptor for substance P), and 5-HT2A receptors would elicit changes in multiple neurochemicals, but would not change breathing. We unilaterally reverse-dialyzed a cocktail of antagonists targeting these receptors into the VRC of intact adult goats. Breathing was continuously monitored while effluent fluid from dialysis was collected for quantification of neurochemicals. We found that neither double blockade of mACh and NK1 receptors, nor triple blockade of mACh, NK1, and 5-HT2A receptors significantly affected breathing (P ≥ 0.05) in goats that were awake or in non-rapid eye movement (NREM) sleep. However, both double and triple blockade increased the effluent concentration of substance P (P < 0.001) and decreased GABA concentrations. These findings support our hypothesis and, together with past data, suggest that both in wakefulness and NREM sleep, multiple neuromodulator systems collaborate to stabilize breathing when a deficit in one or multiple excitatory neuromodulators exists.

Effects on breathing of agonists to μ-opioid or GABA A receptors dialyzed into the ventral respiratory column of awake and sleeping goats

“…This instability contrasts to studies (Langer et al, 2016; Muere et al, 2013, 2015a; Muere et al, 2015b; Muere et al, 2015c) in which dialyses of antagonists to multiple excitatory receptors in the VRC did not destabilize breathing. The destabilization with DAMGO could be due to time-dependent changes in the direct effect of opioid-induced depression and the indirect effect of potential compensatory mechanisms.…”

“…The current and past dialysis studies in unanesthetized goats found neuromodulator compensation during perturbation of G protein-coupled receptors (Muere et al, 2013, 2015a; Muere et al, 2015b; Muere et al, 2015c). Herein we found what could be interpreted as overcompensation for increases in inhibitory ionotropic GABA receptor activity during muscimol dialysis.…”

“…Microdialysis of antagonists to excitatory, neuromodulatory, G protein-coupled receptors into the VRC of awake and sleeping goats (Langer et al, 2016; Muere et al, 2013, 2015a; Muere et al, 2015b; Muere et al, 2015c) support the concept of neuromodulator compensation, whereby decreases in excitatory receptor activity are compensated by increases in other (predominantly excitatory) neuromodulators to prevent a decrease in ventilation and breathing frequency (Doi and Ramirez, 2008, 2010). In other words, there is “compensation” defined as no change in ventilation with dialysis of excitatory antagonists and that this may be due to the observed concomitant increase in excitatory (SP) and decrease in inhibitory neurotransmitters (GABA).…”

“…This secondary/compensatory response may have been relatively greater with 100 μM compared to 50 μM which is why a significant depression in ventilation was found with 50 μM but not 100 μM DAMGO. A secondary/compensatory response is sufficiently rapid to prevent a detectable decrease in ventilation during dialysis of antagonists to excitatory neuromodulator receptors (Muere et al, 2013, 2015a; Muere et al, 2015b; Muere et al, 2015c). Compensation during DAMGO dialysis may also have been within the contralateral VRC.…”

“…Subsequent studies in this goat model found that a lower dose of atropine (5 mM), or selective M2 (methoctramine) or M3 (4-DAMP) receptor antagonists had no effect on f and V̇ I , but increased SP in the effluent dialysate to a lesser extent than 50 mM atropine (Muere et al, 2015a). Moreover, dialysis into the VRC of antagonists to excitatory SP receptors (neurokinin-1; NK-1) or 5-HT receptors (5-HT 2A ) alone or in combination with 5 mM atropine did not affect f and V̇ I , but increased SP and decreased GABA in effluent dialysate (Muere et al, 2015b). These findings suggest that presumptive decreases in activity of one or more excitatory receptors are accompanied by changes in other neuromodulators to maintain f and V̇ I in awake and NREM sleep states which supports the concept of neuromodulator interdependence (Doi and Ramirez, 2008).…”