Optogenetic stimulation of PICo neurons and the respiratory‐sympathetic coupling in in vivoCIH and control mice

Abstract

The resting respiratory activity is generated by synchronized activity of a complex located within the brainstem. This contains a primary oscillator essential for the respiratory rhythmicity and is composed of pacemaker inspiratory neurons. However, recent studies suggest the postinspiratory complex (PiCo) is a novel respiratory oscillator which is critical for the postinspiratory activity in vivo and contains cholinergic and glutamatergic neurons. In addition to the generation of the respiration, studies suggest that the respiratory neurons also interact with the presympathetic neurons involved with the generation of sympathetic activity, contributing to the excitation of sympathetic activity in chronic intermittent hypoxia (CIH) conditions and promoting changes inthe respiratory‐sympathetic coupling. To investigate the contribution of PiCo in the respiratory‐sympathetic coupling, we used an optogenetic in vivo mouse preparation previously exposed to CIH for 21 days (5% O2 – 80 bouts/day). In the end of the protocol, the ventral surface of the brainstem in spontaneously breathing, anaesthetized (Urethane 1.5mg/kg) ChATCre/Ai32 adult and Vglut2cre/Ai32 mice was exposed, and an optical fiber positioned bilaterally over PiCo, while recording the diaphragm muscle (DIA), hypoglossal (HN), cervical vagus (cVN) and cervical sympathetic nerves (cSN). Our data showed that CIH protocol increased the respiratory frequency, post‐I vagus activity and sympathetic activity in the baseline conditions. Random light stimulation (10s) of PiCo ChAT neurons (~ 9mV, 447nm) in significantly increased the cSN discharge during postinspiration in control group. Although the sympathetic activity was increased during all respiratory phases in CIH mice, the stimulation of cholinergic neurons at PiCo did not change the correlation between post‐I and E2 phase. The stimulation of Vglut2 neurons at PiCo area evoked post‐inspiratory discharge of cSN in both groups. Our data demonstrates that during resting conditions, mice exposed to CIH for 3 weeks increased the baseline respiratory frequency and sympathetic activity in vivo and the optogenetic stimulation of PiCo neurons blunts the phasic respiratory –sympathetic coupling in CIH mice, increasing the post‐inspiratory discharge of the sympathetic nerve.