Genetically defined nucleus incertus neurons differ in connectivity and function.

Abstract

The nucleus incertus (NI), a conserved hindbrain structure implicated in the stress response, arousal, and memory, is a major site for production of the neuropeptide relaxin-3. On the basis of <i>goosecoid homeobox 2</i> (<i>gsc2</i>) expression, we identified a neuronal cluster that lies adjacent to <i>relaxin 3a</i> (<i>rln3a</i>) neurons in the zebrafish analogue of the NI. To delineate the characteristics of the <i>gsc2</i> and <i>rln3a</i> NI neurons, we used CRISPR/Cas9 targeted integration to drive gene expression specifically in each neuronal group, and found that they differ in their efferent and afferent connectivity, spontaneous activity, and functional properties. <i>gsc2</i> and <i>rln3a</i> NI neurons have widely divergent projection patterns and innervate distinct subregions of the midbrain interpeduncular nucleus (IPN). Whereas <i>gsc2</i> neurons are activated more robustly by electric shock, <i>rln3a</i> neurons exhibit spontaneous fluctuations in calcium signaling and regulate locomotor activity. Our findings define heterogeneous neurons in the NI and provide new tools to probe its diverse functions.