BHQ-O-5HT Excites Trigeminal Neurons in Intact Zebrafish Larva

(A) Lateral brightfield view of a zebrafish larva at 5 dpf showing placements of the field recording (black arrowhead) and microinjection (open white arrowhead) pipettes relative to the trigeminal ganglion (gV). Dorsal is up and anterior is left, with the eye, hindbrain and ear (Oto, otocyst) indicated for reference. The inset is a confocal fluorescence image of the trigeminal ganglion obtained from a comparable experiment using a 5 dpf larva expressing the cameleon calcium indicator in all neurons. Placements of the recording (arrowhead) and injection (open arrowhead) pipettes are indicated.

(B and C) In vivo extracellular field recordings of 5-HT-induced changes in trigeminal ganglion activity. (B) Baseline recordings from the ventral aspect of the trigeminal ganglion show low-amplitude neural activity. Microinjection (inj.) of 0.5 nl of a 1 mM buffered 5-HT solution in the region of the putative maxillary nerve elicited a brief bust of high-amplitude spiking; in some cases, this initial discharge was followed by a second burst within a few seconds of the injection. Bars above traces denote significant changes from baseline activity. No change in activity was observed upon exposure to three 1 ms pulses of 365 nm light (1PE) spaced <15 s apart. Electronic spikes associated with the lamp discharge are denoted with asterisks; these occur about 60% of the time. (C) Microinjection of 1 nl of a 500 mM buffered BHQ-O-5HT solution did not alter baseline activity. Photolysis of BHQ-O-5HT by exposure to 1 ms pulses of 365 nm light elicited high-amplitude spiking that typically lasted a few seconds (bars). The traces comparing 5-HT to BHQ-O-5HT are temporally aligned to facilitate comparison. A similar experiment was carried out in the optic tectum of 7 dpf zebrafish larva (Figure S2).