Purinergic signaling in cochlear supporting cells reduces hair cell excitability by increasing the extracellular space

Abstract

Neurons in developing sensory pathways exhibit spontaneous bursts of electrical 11 activity that are critical for survival, maturation and circuit refinement. In the auditory system, 12 intrinsically generated activity arises within the cochlea, but the molecular mechanisms that 13 initiate this activity remain poorly understood. We show that burst firing of mouse inner hair cells 14 prior to hearing onset requires P2RY1 autoreceptors expressed by inner supporting cells. P2RY1 15 activation triggers K + efflux and depolarization of hair cells, as well as osmotic shrinkage of 16 supporting cells that dramatically increased the extracellular space and speed of K + 17 redistribution. Pharmacological inhibition or genetic disruption of P2RY1 suppressed neuronal 18 burst firing by reducing K + release, but unexpectedly enhanced their tonic firing, as water 19 resorption by supporting cells reduced the extracellular space, slowing K + clearance. These 20 studies indicate that purinergic signaling in supporting cells regulates hair cell excitability by 21 controlling the volume of the extracellular space. 22 23 Introduction 24The developing nervous system must generate, organize, and refine billions of neurons and their 25 connections. While molecular guidance cues forge globally precise neuronal connections between 26 distant brain areas (Stoeckli, 2018; Dickson, 2002), the organization of local connections is initially 27 coarse and imprecise (Dhande et al., 2011; Kirkby et al., 2013; Sretavan and Shatz, 1986). Coinci-28 dent with the refinement of topographic maps, nascent circuits experience bursts of intrinsically 29 generated activity that emerge before sensory systems are fully functional (Kirkby et al., 2013). 30 This intrinsically generated activity consists of periodic bursts of high frequency firing that pro-31 motes the survival and maturation of neurons in sensory pathways (Blankenship and Feller, 2010; 32 Moody and Bosma, 2005). The precise patterning of this electrical activity appears crucial for re-33 finement of local connections, as its disruption results in improper formation of topographic maps 34 (Antón-Bolaños et al., 2019; Burbridge et al., 2014; Xu et al., 2011) and impaired maturation and 35 specification of sensory neurons (Shrestha et al., 2018; Sun et al., 2018). In all sensory systems 36that have been examined, spontaneous burst firing arises within their respective developing sen-37 sory organs, e.g. retina, olfactory bulb, spindle organ, and cochlea (Blankenship and Feller, 2010). 38 Although the mechanisms that induce spontaneous activity in the developing retina have been ex-39 1 of 26 Manuscript submitted to eLife tensively explored, much less is known about the key steps involved in triggering auditory neuron 40 burst firing in the developing cochlea. Understanding these processes may provide novel insights 41 into the causes of developmental auditory disorders, such as hypersensitivity to sounds and audi-42 tory processing disorders that prevent children from communicating a...

Efferent feedback controls bilateral auditory spontaneous activity

“…Only the dorsal parts of the IC, including DCIC and LCIC, are accessible for calcium imaging 8,23 . Consistent with the previous reports 8,10 , typical spontaneous calcium transients in the dorsal surface of the IC appear as stationary and discrete events that (1) have a relatively homogeneous intensity profile along their major axis, (2) show confined spread along the future tonotopic axis that is approximately perpendicular to the major axis (Fig. 1c and Supplementary Fig.…”

“…In the cochleae, groups of inner hair cells (IHC) fire bursts of action potentials that propagate to central auditory nuclei via ascending pathways to coordinate activity throughout the auditory system prior to hearing onset [5][6][7][8] . Although central auditory activity may inherit many properties directly from the periphery, it manifests with distinct patterns and novel features [8][9][10] . Consequently, understanding the characteristics of spontaneous activity in the central auditory system may provide insight into the maturation of circuit connectivity and the development of higher-order auditory functions.…”

“…Recent studies have shown coordinated spontaneous activity in the inferior colliculi (IC) at P6-P8, utilizing SNAP25-GCaMP6s mice with wide-field epifluorescent microscopy 8,10 . The calcium transients, detected with the GCaMP sensor, manifest as bandshaped bursts that align across the expected future tonotopic axis in the IC.…”

Pregnancy-associated plasma protein-aa regulates endoplasmic reticulum–mitochondria associations

“…We show that Pappaa is essential for the function of ER and its relationship to mitochondria in hair cells. Hair cell function and activity are influenced by the surrounding support cells as well as the efferent and afferent neurons that innervate hair cells (Babola et al, 2020;Faucherre et al, 2009;Pichler and Lagnado, 2020). A caveat to our pharmacological experiments is that drug treatments will affect all of these cell types, and some effects on hair cells may be indirect.…”

BAC transgenic mice to study the expression of P2X2 and P2Y1 receptors

“…Additionally, a P2RY1 knockout with lacZ reporter (P2RY1 tm1(KOMP)Vlcg ) produced by Regeneron Pharmaceuticals (Tarrytown, NY, USA) for the knockout mouse project (KOMP) at the University of California is meanwhile available and was recently used to study P2Y 1 R expression in the developing cochlea [51]. It was shown that burst firing of mouse inner hair cells prior to hearing onset requires P2RY1 autoreceptors expressed by inner supporting cells and that purinergic signaling in cochlear supporting cells regulates hair cell excitability by volume control of the extracellular space [51].…”