Maintaining stable body temperature through environmental thermal stressors requires detection of temperature changes, relay of information, and coordination of physiological and behavioral responses. Studies have implicated areas in the preoptic area of the hypothalamus (POA) and the parabrachial nucleus (PBN) as nodes in the thermosensory neural circuitry and indicate that the opioid system within the POA is vital in regulating body temperature. In the present study we identify neurons projecting to the POA from PBN expressing the opioid peptides dynorphin and enkephalin. Using mouse models, we determine that warm-activated PBN neuronal populations overlap with both prodynorphin (Pdyn) and proenkephalin (Penk) expressing PBN populations. Here we report that in the PBN Prodynorphin (Pdyn) and Proenkephalin (Penk) mRNA expressing neurons are partially overlapping subsets of a glutamatergic population expressing Solute carrier family 17 (Slc17a6) (VGLUT2). Using optogenetic approaches we selectively activate projections in the POA from PBN Pdyn, Penk, and VGLUT2 expressing neurons. Our findings demonstrate that Pdyn, Penk, and VGLUT2 expressing PBN neurons are critical for physiological and behavioral heat defense.

Maintaining stable body temperature through environmental thermal stressors requires detection of temperature changes, relay of information, and coordination of physiological and behavioral responses. Studies have implicated areas in the preoptic hypothalamic area (POA) and the parabrachial nucleus (PBN) as nodes in the thermosensory neural circuitry and indicate the opioid system within the POA is vital in regulating body temperate. In the present study we identify neurons projecting to the POA from PBN expressing the opioid peptides Dynorphin (Dyn) and Enkephalin (Enk). We determine that warm-activated PBN neuronal populations overlap with both Dyn and Enk expressing PBN populations. We demonstrate that Dyn and Enk expressing neurons are partially overlapping subsets of a glutamatergic population in the PBN. Using optogenetic approaches we selectively activate projections in the POA from PBN Dyn, Enk, and VGLUT2 expressing neurons. Our findings demonstrate that Dyn, Enk, and VGLUT2 expressing PBN neurons are critical for physiological and behavioral heat defense. potential roles of opioid receptor mediated behaviors in both Dyn+ and Enk+ PBN-POA projections. Here we report that glutamatergic, Dyn+, and Enk+ neuronal populations projecting from PBN to POA initiate physiological and behavioral heat defensive behaviors. Chemogenetic inhibition of glutamatergic PBN neurons blocks vasomotor responses to thermal heat challenge.The studies reported here provide new insights into the thermoregulatory properties of parabrachial neuropeptide-containing projections to the hypothalamus in homeostatic and metabolic behavior. Results Ambient warmth activates Dyn+ and Enk+ neurons in PBNEffects of mu and kappa receptor signaling on body temperature have been described and mRNA for preprodynorphin and preproenkephalin has been reported to be expressed in the PBN (Baker and Meert, 2002;Chen et al., 2005;Clark, 1979;Engstrom et al., 2001;Hermanson and Blomqvist, 1997;Hermanson et al., 1998). To examine if PBN neurons expressing dynophin or enkephalin opioid neuropeptides are activated by ambient warmth, we exposed mice to ambient warmth (38°C) or room temperature (21-23°C) for four hours prior to preparation of brain for fos staining. We performed immunohistochemistry (IHC) on collected brains sections containing the PBN with antibody directed against c-Fos (anti-c-Fos) to examine induction of c-Fos expression as a marker of neuronal activation (Sheng and Greenberg, 1990). Consistent with recent reports, we observed induction of c-Fos expression in the lateral PBN (LPBN) (Figure 1B-C) (Geerling et al., 2016). In brain sections from warm exposed mice (n=8) compared to room temperature controls (n=4), c-Fos staining revealed a robust and significant (p=0.003) increase in mean ±SEM number of neurons positive for c-Fos expression in the LPBN per brain: 265.8 ±41.9 in warm exposed mice compared to 23.2 ±4.0 in room temperature controls (Figure 1C).Cells in LPBN, lateral to superior cerebellar peduncle, in sections corresponding -5.0 ...

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