Modulation of bulbospinal rostral ventral lateral medulla neurons by hypoxia/hypercapnia but not medullary respiratory activity

Abstract

Although sympathetic vasomotor discharge has respiratory modulation, the site(s) responsible for this cardiorespiratory interaction is unknown. One likely source for this coupling is the rostral ventral lateral medulla (RVLM), where presympathetic neurons originate in close apposition to respiratory neurons. The current study tested the hypothesis that RVLM bulbospinal neurons are modulated by medullary respiratory network activity using whole-cell patch-clamp electrophysiological recordings of RVLM neurons while simultaneously recording fictive respiratory bursting activity from the hypoglossal rootlet. Additionally, we examined whether challenges to cardiorespiratory function, mainly hypoxia/hypercapnia, alter the activity of bulbospinal neurons and, secondarily, whether changes in synaptic input mediate these responses. Surprisingly, our results indicate that inspiratory-related activity did not modulate glutamatergic, γ-aminobutyric acid-ergic, or glycinergic synaptic events or spontaneous action potential firing in these RVLM neurons. However, hypoxia/hypercapnia reversibly decreased the frequency of γ-aminobutyric acid and glycine inhibitory postsynaptic currents. Glycinergic inhibitory postsynaptic current frequency was depressed from the fifth through the 10th minute, whereas the depression of γ-aminobutyric acid-ergic events became significant only at the 10th minute of hypoxia/hypercapnia. On the basis of spontaneous firing activity, there were 2 populations of RVLM bulbospinal neurons. The firing frequency of low-discharging RVLM neurons was facilitated by hypoxia/hypercapnia, and this increase depended on reduced inhibitory neurotransmission. The firing frequency in RVLM neurons with high-discharge rates was inhibited, independent of synaptic input, by hypoxia/hypercapnia. This article demonstrates that sympathetic-respiratory coupling is not active in the neonatal brain stem slice, and reductions in inhibitory neurotransmission to low spontaneously active bulbospinal RVLM neurons are responsible for hypoxia/hypercapnia-elicited increases in activity.

Figure 1. RVLM bulbospinal neurons not demonstrate spontaneous inspiratory-like modulation in glutamergic (n=7), GABAergic (n=7), or glycinergic (n=7) postsynaptic current (PSC) frequencies during normoxic conditions

Inspiratory-like bursting activity was recorded from the hypoglossal (XII) rootlet and electronically integrated (fXII). Labeled RVLM neurons were patch-clamped in the whole-cell configuration and glutamatergic, GABAergic and glycinergic PSC were individually isolated and recorded. Arrows indicates where inspiratory-like bursts occur.

Figure 2. During hypoxia/hypercapnia, spontaneous inhibitory postsynaptic current (IPSC) frequencies were decreased in RVLM bulbospinal neurons

GABAergic frequencies (n=7) became significantly depressed from control (Pre) conditions only by the tenth minute of hypoxia/hypercapnia (top panel; p<0.05). Glycinergic frequencies (n=7) became significantly lower from Pre at the fifth minute and this depression was maintained through the tenth minute of hypoxia/hypercapnia (top panel; p<0.05). * indicates significant differences (p<0.05) from Pre.

Figure 3. RVLM bulbospinal neurons demonstrated opposing responses to hypoxia/hypercapnia depending on their spontaneous action potential (AP) frequencies

Slow-firing RVLM neurons (n=7) significantly increased AP frequencies at the fifth and tenth minute of hypoxia/hypercapnia (top panel; p<0.05); whereas the fast-firing subtype (n=6) significantly decreased their AP frequencies only at the tenth minute of hypoxia/hypercapnia (bottom panel; p<0.05). * indicates significant differences (p<0.05) from Pre.

Figure 4. The increase in spontaneous action potential (AP) frequencies during hypoxia/hypercapnia seen in slow-firing RVLM bulbospinal neurons (n=7) was attenuated by inhibitory neurotransmission blockade

After focal application of gabazine and strychnine to block inhibitory neurotransmission, slow-firing RVLM neurons (n=7) did not significantly increase AP frequencies at any time during hypoxia/hypercapnia (top panel; p<0.05) compared to the control period (Pre); Additionally, in the fast-firing subtype (n=6) no changes in AP frequency were seen during hypoxia/hypercapnia in the presence of gabazine and strychnine (bottom panel; p>0.05).