RESUMEN

RFamide neuropeptide VF (NPVF) is expressed by neurons in the hypothalamus and has been implicated in nociception, but the circuit mechanisms remain unexplored. Here, we studied the structural and functional connections from NPVF neurons to downstream targets in the context of nociception, using novel transgenic lines, optogenetics, and calcium imaging in behaving larval zebrafish. We found a specific projection from NPVF neurons to serotonergic neurons in the ventral raphe nucleus (vRN). We showed NPVF neurons and vRN are suppressed and excited by noxious stimuli, respectively. We combined optogenetics with calcium imaging and pharmacology to demonstrate that stimulation of NPVF cells suppresses neuronal activity in vRN. During noxious stimuli, serotonergic neurons activation was due to a suppression of an inhibitory NPVF-ventral raphe peptidergic projection. This study reveals a novel NPVF-vRN functional circuit modulated by noxious stimuli in vertebrates.

Asunto(s)

Hipotálamo/metabolismo , Neuropéptidos/metabolismo , Nocicepción , Núcleos del Rafe/metabolismo , Pez Cebra/metabolismo , Secuencia de Aminoácidos , Animales , Neuronas/metabolismo , Neuropéptidos/química , Serotonina/metabolismo

RESUMEN

Neurons exhibit rhythmic activity that ultimately affects behavior such as sleep. In living zebrafish larvae, we used time-lapse two-photon imaging of the presynaptic marker synaptophysin in hypocretin/orexin (HCRT) neurons to determine the dynamics of synaptic modifications during the day and night. We observed circadian rhythmicity in synapse number in HCRT axons. This rhythm is regulated primarily by the circadian clock but is also affected by sleep deprivation. Furthermore, NPTX2, a protein implicated in AMPA receptor clustering, modulates circadian synaptic changes. In zebrafish, nptx2b is a rhythmic gene that is mostly expressed in hypothalamic and pineal gland cells. Arrhythmic transgenic nptx2b overexpression (hcrt:NPTX2b) increases synapse number and abolishes rhythmicity in HCRT axons. Finally, hcrt:NPTX2b fish are resistant to the sleep-promoting effects of melatonin. This behavioral effect is consistent with NPTX2b-mediated increased activity of HCRT circuitry. These data provide real-time in vivo evidence of circadian and homeostatic regulation of structural synaptic plasticity.

Asunto(s)

Ritmo Circadiano/fisiología , Homeostasis/fisiología , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Plasticidad Neuronal/fisiología , Neuronas/fisiología , Neuropéptidos/metabolismo , Sinapsis/fisiología , Animales , Animales Modificados Genéticamente , Axones/metabolismo , Conducta Animal , Encéfalo/citología , Encéfalo/crecimiento & desarrollo , Proteína C-Reactiva/genética , Proteína C-Reactiva/metabolismo , Ritmo Circadiano/genética , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas Fluorescentes Verdes/genética , Homeostasis/genética , Humanos , Técnicas In Vitro , Larva , Luz , Melatonina/farmacología , Microscopía Confocal , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Plasticidad Neuronal/genética , Neuronas/citología , Orexinas , Glándula Pineal/crecimiento & desarrollo , Glándula Pineal/metabolismo , Sinapsis/genética , Sinaptofisina/metabolismo , Pez Cebra