The Transfer Characteristics of Hair Cells Encoding Mechanical Stimuli in the Lateral Line of Zebrafish

Abstract

Hair cells transmit mechanical information by converting deflection of the hair bundle into synaptic release of glutamate. We have investigated this process in the lateral line of larval zebrafish (male and female) to understand how stimuli are encoded within a neuromast. Using multiphoton microscopy in vivo, we imaged synaptic release of glutamate using the reporter iGluSnFR as well as deflections of the cupula. We found that the neuromast is composed of a functionally diverse population of hair cells. Half the hair cells signaled cupula motion in both directions from rest, either by increasing glutamate release in response to a deflection in the positive direction or by reducing release in the negative direction. The relationship between cupula deflection and glutamate release demonstrated maximum sensitivity at displacements of just ∼40 nm in the positive direction. The remaining hair cells only signaled motion in one direction and were less sensitive, extending the operating range of the neuromast beyond 1 μm. Adaptation of the synaptic output was also heterogeneous, with some hair cells generating sustained glutamate release in response to a steady deflection of the cupula and others generating transient outputs. Finally, a distinct signal encoded a return of the cupula to rest: a large and transient burst of glutamate release from hair cells unresponsive to the initial stimulus. A population of hair cells with these different sensitivities, operating ranges, and adaptive properties will allow the neuromast to encode weak stimuli while maintaining the dynamic range to signal the amplitude and duration of stronger deflections.SIGNIFICANCE STATEMENT Hair cells transmit information about mechanical stimuli by converting very small deflections of their hair bundle into changes in the release of the neurotransmitter glutamate. We have measured this input/output relation in the live fish using a fluorescent protein and find that different hair cells vary in their mechanical sensitivity and the time course of their response. These variations will allow the fish to sense the timing and duration of both very weak stimuli (∼40 nm deflections) and strong stimuli (∼1 μm), underlying the ability of the fish to avoid predators and maintain its body position in flowing water.

Motor behaviour selectively inhibits hair cells activated by forward motion in the lateral line of Zebrafish

“…Motor nerve activity was measured electrophysiologically while optical reporters were targeted genetically to measure calcium activity in efferent and/or afferent neurons as well as glutamate release from the synapses of hair cells in neuromasts towards the back of the tail (the posterior lateral line). These various measurements were combined with the application of mechanical stimuli to individual neuromasts to assess changes in sensitivity (Fig 1A -D; Pichler and Lagnado, 2019).…”

“…To what extent does the efference copy modulate the output from a neuromast? To investigate this question we monitored the synaptic output from hair cells by expressing the glutamate sensor iGluSnFR (Marvin et al, 2013) under the control of the Sill promoter (Pichler and Lagnado, 2019;Pujol-Marti and Lopez-Schier, 2013); Fig 1B & C and Fig 2A & B). In these experiments we did not paralyze fish by the usual method of applying the neuromuscular blocker α-BTX because this agent has also been reported to block the α9/α10 isoforms of nicotinic acetylcholine receptors (nAChR) present in hair cells (Erickson and Nicolson, 2015;Verbitsky et al, 2000).…”

“…Similar suppression of spontaneous activity was observed in four neuromasts out of eight, and could also be observed by monitoring glutamate release at the output of afferent neurons in the Medial Octavolateralis Nucleus (MON) (Fig S3). Synaptic activity in the absence of a mechanical stimulus is a key aspect of the "push-pull" system by which a population of hair cells of opposite polarity signals stimulus direction (Pichler and Lagnado, 2019) and blocking spontaneous release will prevent the signalling of direction by the hair cells inhibited by a particular direction of motion.…”

“…Neuromasts also receive inputs from cholinergic efferents that modulate the sensory signal transmitted to the hindbrain (Bricaud et al, 2001;Chagnaud et al, 2015;Flock and Russell, 1976). The mechanical sensitivity of hair cells varies, but some detect displacements of just a few tens of nanometers (Pichler and Lagnado, 2019).…”

“…First, it will protect the lateral system from overstimulation (Roberts, 1989). In the absence of efferent modulation, swimming would generate a strong stimulus that would induce adaptation by, for instance, depletion of synaptic vesicles at the ribbon synapse of hair cells (Goutman, 2017;Pichler and Lagnado, 2019;Schnee et al, 2011;Schnee et al, 2005). At the ribbon synapse of auditory hair cells, recovery from synaptic depression is only complete after seconds of rest (Cho et al, 2011), which would introduce a "dead-time" at the end of a swimming bout when the lateral line was not signaling effectively.…”