Analyzing the brainstem circuits for respiratory chemosensitivity in freely moving mice

Abstract

Regulation of systemic PCO2 is a life-preserving homeostatic mechanism. In the medulla oblongata, the retrotrapezoid nucleus (RTN) and rostral medullary Raphe are proposed as CO2 chemosensory nuclei mediating adaptive respiratory changes. Hypercapnia also induces active expiration, an adaptive change thought to be controlled by the lateral parafacial region (pFL). Here we use GCaMP6 expression and head-mounted mini-microscopes to image Ca2+ activity in these nuclei in awake adult mice during hypercapnia. Activity in the pFL supports its role as a homogenous neuronal population that drives active expiration. Our data show that chemosensory responses in the RTN and Raphe differ in their temporal characteristics and sensitivity to CO2, raising the possibility these nuclei act in a coordinated way to generate adaptive ventilatory responses to hypercapnia. Our analysis revises the understanding of chemosensory control in awake adult mouse and paves the way to understanding how breathing is coordinated with complex non-ventilatory behaviours.

Whole-brain analysis of CO2chemosensitive regions and identification of the retrotrapezoid and medullary raphé nuclei in the common marmoset (Callithrix jacchus)

“…Aside from the Phox2b + neuronal population in the RTN, the 5-HT neurons of the medullary 10aphe are also proposed to act as central CO 2 chemosensor in rodents (Richerson et al 2001; Richerson 2004; Nattie et al 2004; Hodges and Richerson 2010; Corcoran et al 2009; Ray et al 2011; Bhandare et al 2020). Similar to data in rodents, we found that a subset of serotonergic neurons in the brainstem are activated by systemic hypercapnia (Figure 3).…”

Neural correlate of reduced respiratory chemosensitivity during chronic epilepsy

“…Using methods that we have recently developed (Bhandare et al, 2022 ), neurons of the RTN and RVLM were transduced with an AAV to drive expression of Ca 2+ reporter GCaMP6 under the control of the hSyn promoter and to allow the assessment of their activity via a head-mounted mini-microscope ( Figure 1A ). We prepared 18 mice (9 with an injection of GCaMP6 into the RTN and 9 with injection into the RVLM, Figure 1B ) and assessed their responses to hypercapnia before and weeks after induction of SE (during the chronic phase of epilepsy; 3, 5, and 7 weeks post-SE).…”

“…Detailed procedures to assess and compensate for movement artifacts have been presented in previously studies (Bhandare et al, 2022 ). In brief, if there was too much uncompensated motion artifact so that we could not clearly analyse Ca 2+ signals, we excluded these recordings from quantitative analysis.…”

“…Cardiovascular function is controlled by excitatory rostral ventrolateral medullary (RVLM) neurons in the brainstem (Spyer and Gourine, 2009 ; Guyenet, 2014 ). To address whether progressive changes in epilepsy alter the neuronal firing patterns and chemosensitivity in these critical brainstem networks, we used intravital microscopy to record these brainstem nuclei in freely behaving mice (Bhandare et al, 2022 ).…”

Shining the Light on Astrocytic Ensembles

“…In addition, astrocytes play an important role in intercellular information transmission through neuronal oscillations, which contribute to rhythmicity. They are essential participants in understanding the synchronous activities of central pattern generators (CPGs), which control behaviors such as respiration [40][41][42][43][44], locomotion [45][46][47][48][49], mastication [50][51][52], and the regulation of sleep patterns [53]. Astrocytes in the suprachiasmatic nucleus of the hypothalamus have been shown to independently encode circadian information and control the period of circadian oscillations through glutamate [54][55][56], and the inhibition of vesicular release in hippocampal astrocytes provokes disruption of gamma oscillatory activity [57].…”