Activity-independent specification of synaptic targets in the posterior lateral line of the larval zebrafish

Abstract

The development of functional neural circuits requires that connections between neurons be established in a precise manner. The mechanisms by which complex nervous systems perform this daunting task remain largely unknown. In the posterior lateral line of larval zebrafish, each afferent neuron forms synaptic contacts with hair cells of a common hair-bundle polarity. We investigated whether afferent neurons distinguish hair-cell polarities by analyzing differences in the synaptic signaling between oppositely polarized hair cells. By examining two mutant zebrafish lines with defects in mechanoelectrical transduction, and by blocking transduction during the development of wild-type fish, we found that afferent neurons could form specific synapses in the absence of stimulus-evoked patterns of synaptic release. Asking next whether this specificity arises through intrinsically generated patterns of synaptic release, we found that the polarity preference persisted in two mutant lines lacking essential synaptic proteins. These results indicate that lateral-line afferent neurons do not require synaptic activity to distinguish hair-cell polarities and suggest that molecular labels of hair-cell polarity guide prepatterned afferents to form the appropriate synapses.calcium channel ͉ hair cell ͉ neuromast ͉ planar cell polarity ͉ protocadherin A n essential feature of neural development is the establishment of specific synaptic connections. To form the appropriate contacts, each growing axon must respond to guidance cues, find its target region, and then establish synapses with specific target cells (1, 2). The first two of these steps-axonal guidance and target recognition-rely predominantly on molecular signposts that attract or repulse growth cones in a manner independent of neuronal activity (3, 4). How neurons decide to form stable synapses with particular target cells, however, remains unclear. Activity serves an important role in regulating the growth of axonal arbors and in selectively stabilizing synapses (5-8). In several vertebrate systems, axons form synapses diffusely within the target region and then undergo activitydependent pruning to eliminate inappropriate synapses (9-14). Hebb's postulate, by which correlated activity between synaptic partners strengthens connections (15,16), offers an attractive model to explain this phenomenon (17). Nevertheless, the evidence for an activity-dependent process must be reconciled with data suggesting that normal brain architecture can form in the absence of synaptic transmission (18)(19)(20). In this case, synaptic specificity could derive from a combinatorial code of cell-surface molecules such as cadherins (21) or members of the immunoglobin superfamily (22). These fundamental uncertainties highlight the need for in vivo studies in an experimentally tractable vertebrate system.The posterior lateral line of zebrafish permits a detailed analysis of the role of activity in establishing synaptic specificity. The larval posterior lateral line consists of superficial clusters ...

Cellular projections from sensory hair cells form polarity-specific scaffolds during synaptogenesis

“…Finally, the extension of projections toward nearby afferent terminals reduces a terminal's search for new synaptic sites from three dimensions to only one. Because polarity-specific innervation of lateral line hair cells occurs in the absence of hair cell activity, a chemoaffinity mechanism may be responsible for partner matching (Nagiel et al 2009). Although hair cell projections are more likely to extend into the vicinity of mature hair cells of the same polarity, this predilection alone is unlikely to confer specific innervation inasmuch as afferent terminals of both functional polarities reside beneath mature hair cells.…”

“…From the posterior lateral line ganglion, which lies immediately caudal to the ear, several bipolar neurons extend sensory axons to contact a neuromast (Metcalfe et al 1985). Each afferent fiber receives synaptic input from hair cells of only one polarity, a specificity that occurs independently of hair cell activity (Nagiel et al 2008(Nagiel et al , 2009). To learn how this synaptic pattern arises, we studied synapse formation by newly differentiated hair cells in larvae 2-4 d post-fertilization (dpf).…”

Ribeye is required for presynaptic CaV1.3a channel localization and afferent innervation of sensory hair cells

“…A recent study examined afferent innervation of lateral line neuromasts in zebrafish mutants lacking vesicular glutamate transporter 3 (Vglut3), the transporter responsible for filling hair cell synaptic vesicles with glutamate (Nagiel et al, 2009;Obholzer et al, 2008). They reported no significant difference in the mean fraction of hair cells innervated by a single afferent neuron in vglut3 mutants verses wild type, and concluded that synaptic transmission did not appear to be essential for synaptic maintenance (Nagiel et al, 2009). These results suggest that Ribeye is probably playing a crucial structural role, perhaps by recruiting additional molecules to stabilize synaptic connections.…”

Early Development of Hearing in Zebrafish

“…The agarose covering the right side of the larva was scraped away with a 31.5-gauge syringe needle so that the right inner ear was exposed for positioning the stimulus probe and recording electrode. The top of the larva was then covered with a 0.25-ml solution containing 116 mM NaCl, 2.9 mM KCl, 1.8 mM CaCl 2 , and 5.0 mM Hepes (pH=7.2;Nagiel et al 2009) to prevent the agarose from drying. The recording chamber was placed in a Petri dish holder that was temperature controlled at ∼28.5°C (Model TC-344B, Warner Instruments, Hamden, CT).…”