Ventrolateral Medullary Functional Connectivity and the Respiratory and Central Chemoreceptor-Evoked Modulation of Retrotrapezoid-Parafacial Neurons

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Bötzinger, breathing, cross-correlation, brainstem, circuits

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The medullary ventral respiratory column (VRC) of neurons is essential for respiratory motor pattern generation; however, the functional connections among these cells are not well understood. A rostral extension of the VRC, including the retrotrapezoid nucleus/parafacial region (RTN-pF), contains neurons responsive to local perturbations of CO2/pH. We addressed the hypothesis that both local RTN-pF interactions and functional connections from more caudal VRC compartments—extending from the Bötzinger and pre-Bötzinger complexes to the ventral respiratory group (Böt-VRG)—influence the respiratory modulation of RTN-pF neurons and their responses to central chemoreceptor and baroreflex activation. Spike trains from 294 RTN-pF and 490 Böt-VRG neurons were monitored with multielectrode arrays along with phrenic nerve activity in 14 decerebrate, vagotomized cats. Overall, 214 RTN-pF and 398 Böt-VRG neurons were respiratory modulated; 124 and 95, respectively, were cardiac modulated. Subsets of these neurons were tested with sequential, selective, transient stimulation of central chemoreceptors and arterial baroreceptors; each cell's response was evaluated and categorized according to the change in firing rate (if any) following the stimulus. Cross-correlation analysis was applied to 2,884 RTN-pF↔RTN-pF and 8,490 Böt-VRG↔RTN-pF neuron pairs. In total, 174 RTN-pF neurons (59.5%) had significant features in short-time scale correlations with other RTN-pF neurons. Of these, 49 neurons triggered cross-correlograms with offset peaks or troughs (n = 99) indicative of paucisynaptic excitation or inhibition of the target. Forty-nine Böt-VRG neurons (10.0%) were triggers in 74 Böt-VRG→RTN-pF correlograms with offset features, suggesting that Böt-VRG trigger neurons influence RTN-pF target neurons. The results support the hypothesis that local RTN-pF neuron interactions and inputs from Böt-VRG neurons jointly contribute to respiratory modulation of RTN-pF neuronal discharge patterns and promotion or limitation of their responses to central chemoreceptor and baroreceptor stimulation.

contemporary views consider the region of the retrotrapezoid nucleus (RTN)/parafacial-area (RTN-pF) to be a rostral extension of the classically defined ventral respiratory column (VRC), which encompasses the Bötzinger (Böt) and pre-Bötzinger (pre-Böt) complexes along with the rostral and caudal ventral respiratory groups (collectively referred to as Böt-VRG). These compartments are key components of the neural network responsible for the generation and modulation of the respiratory rhythm (Abdala et al. 2009a; Alheid et al. 2002; Feldman and Del Negro 2006; Molkov et al. 2010; Onimaru and Homma 2006; Rybak et al. 2007; Smith et al. 1991, 2007).

Some RTN neurons have respiratory-modulated firing patterns (Connelly et al. 1990; Pearce et al. 1989). Anatomical projections to the RTN-pF from medullary areas involved in respiratory control have been demonstrated (Smith et al. 1989). Lesions within the Böt-VRG reduce or eliminate the respiratory modulation of a subset of RTN-region neurons, an effect that has been attributed to loss of inhibitory influences from the respiratory central pattern generator (Guyenet et al. 2005a).

Several lines of evidence support the hypothesis that the RTN-pF is an essential site of central chemoreception (Guyenet 2008; Loeschcke 1982): 1) the region contains chemoresponsive neurons (Nattie et al. 1993); 2) lesions of neurons in the region reduce or eliminate phrenic activity, alter cycle frequency (St. John et al. 1989), and decrease CO2 sensitivity (Nattie et al. 1991); 3) RTN neurons have properties indicative of intrinsic chemosensitivity (Guyenet et al. 2005b; Mulkey et al. 2004; Nattie and Li 1995; Takakura et al. 2006); 4) RTN neurons respond to ATP released by nearby chemosensitive astrocytes (Gourine et al. 2010); and 5) specific deletion of glutamatergic Phox2b-expressing neurons in the RTN-pF region causes a lack of CO2 sensitivity and fatal central apnea (Dubreuil et al. 2008). RTN neurons influenced by both central and peripheral chemoreceptor inputs have also been identified (Takakura et al. 2006); however, pathways contributing to these neuronal response profiles are not known. Functional inputs, whether locally within the RTN-pF region or coming from Böt-VRG neurons, may contribute to the network mechanisms for respiratory modulation and the mediation of chemoresponsiveness. These potential circuit routes remain largely unexplored.

The present study was motivated by the aforementioned gaps in knowledge and sought evidence for the hypothesis that both local RTN-pF neuron interactions and influences from the Böt-VRG contribute to respiratory modulation of RTN-pF region neurons and affect or mediate their central chemoreceptor responses. Identification of the functional connectivity of the respiratory network will enhance our understanding of circuit level mechanisms responsible for the regulation of breathing and its dynamic coordination with other autonomic activities.

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Citation / Publisher Attribution

Journal of Neurophysiology, v. 105, issue 6, p. 2960-2975