Patch Clamp Recordings from Embryonic Zebrafish Mauthner Cells

Abstract

Mauthner cells (M-cells) are large reticulospinal neurons located in the hindbrain of teleost fish. They are key neurons involved in a characteristic behavior known as the C-start or escape response that occurs when the organism perceives a threat. The M-cell has been extensively studied in adult goldfish where it has been shown to receive a wide range of excitatory, inhibitory and neuromodulatory signals 1 . We have been examining M-cell activity in embryonic zebrafish in order to study aspects of synaptic development in a vertebrate preparation. In the late 1990s Ali and colleagues developed a preparation for patch clamp recording from M-cells in zebrafish embryos, in which the CNS was largely intact 2,3,4 . The objective at that time was to record synaptic activity from hindbrain neurons, spinal cord neurons and trunk skeletal muscle while maintaining functional synaptic connections within an intact brain-spinal cord preparation. This preparation is still used in our laboratory today. To examine the mechanisms underlying developmental synaptic plasticity, we record excitatory (AMPA and NMDA-mediated) 5,6 and inhibitory (GABA and glycine) synaptic currents from developing M-cells. Importantly, this unique preparation allows us to return to the same cell (M-cell) from preparation to preparation to carefully examine synaptic plasticity and neuro-development in an embryonic organism. The benefits provided by this preparation include 1) intact, functional synaptic connections onto the M-cell, 2) relatively inexpensive preparations, 3) a large supply of readily available embryos 4) the ability to return to the same cell type (i.e. M-cell) in every preparation, so that synaptic development at the level of an individual cell can be examined from fish to fish, and 5) imaging of whole preparations due to the transparent nature of the embryos.

Central nervous system hypomyelination disrupts axonal conduction and behaviour in larval zebrafish

“…Intriguingly, RS axons are first to be myelinated in the zebrafish CNS and exhibit activity-regulated myelination at larval stages (16,21,22), implying that regulation of their myelination might influence circuit function in early larvae. In vivo electrophysiological recordings from subsets of individual RS neurons are feasible (23)(24)(25), which in principle permits direct measurement of myelinated axon conduction properties underlying behaviour. However, how disruption to CNS myelination affects the behaviour or axonal conduction properties of larval zebrafish remains to be investigated.…”

NMDA receptors on zebrafish Mauthner cells require CaMKII‐α for normal development

“…All procedures were performed in compliance with the guidelines stipulated by the Canadian Council for Animal Care and the University of Alberta. Zebrafish embryos at 2 dpf were dissected and prepared for electrophysiology as described previously (Drapeau et al, ; Roy and Ali, ). In brief, the embryos were anesthetized in 0.02% tricaine (MS‐222) (Sigma‐Aldrich, St Louis, MO) and pinned through notochord onto an electrophysiological recording chamber.…”

The brain as a dynamically active organ

“…Perhaps the most well-known and beststudied escape response is the C-start response in teleost fish. Mediated by the equally wellknown Mauthner cell, the largest vertebrate neuron, it heads the fish away from a threatening stimulus [81][82][83][84][85][86][87][88][89][90][91][92][93][94][95][96]. Sensory neurons in the inner ear or the side-line organ synapse onto the Mauthner neuron which leads to contraction of the contralateral trunk muscles and inhibition of the ipsilateral ones to bend the animal into a Cshaped form with the head pointing away from the stimulus (Fig.…”