Substance P modulation of TRPC3/ 7 channels improves respiratory rhythm regularity and ICAN‐dependent pacemaker activity

Abstract

Neuromodulators, such as Substance P (SubP) play an important role in modulating many rhythmic activities driven by central pattern generators (e.g., locomotion, respiration). However, the mechanism by which SubP enhances breathing regularity has not been determined. Here, we used mouse brainstem slices containing the pre-Bötzinger Complex (Pre-BötC) to demonstrate, for the first time, that SubP activates transient receptor protein canonical (TRPC) channels to enhance respiratory rhythm regularity. Moreover, SubP enhancement of network regularity is accomplished via selective enhancement of ICAN-dependent intrinsic bursting properties. In contrast to INaPdependant pacemakers, ICAN-dependant pacemaker bursting activity is TRPC dependent. Western Blots reveal TRPC3 and TRPC7 channels are expressed in rhythmically active ventral respiratory group (VRG) island preparations. Taken together, these data suggest that SubPmediated activation of TRPC3/7 channels underlies rhythmic ICAN-dependent pacemaker activity and enhances the regularity of respiratory rhythm activity.

The role of spiking and bursting pacemakers in the neuronal control of breathing

“…SP has been shown to activate neurokinin-1 receptors [100] that are coupled to Gq/11 proteins, that in turn activate phospholipase C. This intracellular cascade eventually induces I CAN , a mechanism that likely contributes to the effects of SP on burst amplitude. Recently, it has been suggested that this mechanism involves the activation of transient receptor protein canonical channels (TRPCs) [75], a mechanism which was also demonstrated in HEK293 cells [104]. SP-mediated effects on respiratory neurons are reversed by the application of an I CAN antagonist (flufenamic acid), but not by an I NaP antagonist (riluzole) in vitro.…”

“…However, despite this circumstantial evidence, the molecular identity of the I CAN current remains unresolved. Ben-Mabrouk and Tryba [75] could not verify TRPM4/5 expression in the preBötC and proposed evidence that I CAN may instead depend on the TRPC3 or TRPC7 channels. In their experiments, the authors used the antagonist SKF-96365 to show that the effect of NK-1 receptor activation by substance P is reversible by blockade of the TRPC channels and that they could reverse the effects of substance P on CS pacemaker neurons.…”

“…SP-mediated effects on respiratory neurons are reversed by the application of an I CAN antagonist (flufenamic acid), but not by an I NaP antagonist (riluzole) in vitro. Since TRPCs are non-selective cation channels permeable to monovalent and divalent ions [105], they might mediate I CAN in CS neurons [75]. Thus, similar to NE, SP modulates pacemaker properties in respiratory neurons via I CAN and it can alter the balance between autonomous spiking, bursting, and synaptic mechanisms.…”

“…Substance P activates the inspiratory frequency at the network and behavioral level [9,32,93,[99][100][101][102] and also plays a critical role in the stability of the respiratory rhythm in vitro [75,93,103] and in vivo [99]. At the cellular level, SP slowly depolarizes spiking pacemaker neurons, leading to an increase in their intrinsic firing rate.…”

“…The channels that mediate the I CAN have been suggested to belong to the transient receptor potential (TRP) family [75,76]. These channels were first discovered in Drosophila [77] and consist of six transmembrane spanning segments (S1-S6) with a cation permeable pore (S5-S6).…”