Long-range circuit of the mouse spinal trigeminal nucleus caudalis

Background The spinal trigeminal nucleus caudalis (SPVC) is a key central relay in the trigeminal circuit that processes orofacial nociceptive and somatosensory information. However, its long-range connectivity patterns remain poorly characterized. Methods Using viral tracing and fluorescence micro-optical sectioning tomography (fMOST), we obtained three-dimensional datasets of long-range input-output circuits of the SPVC in adult male C57BL/6J mice. An orofacial pain model was established by injecting formalin into the whisker pad of adult male C57BL/6J mice, followed by pain behavior assays and cfos immunohistochemical staining. Using adult male Fos-CreER mice combined with viral tracing and fMOST imaging, we mapped the projection patterns of pain-activated neurons in the SPVC. Results We systematically mapped the long-range connectivity patterns of SPVC in adult male mice. High-resolution, three-dimensional whole-head imaging with the fMOST system revealed substantial neuronal input to the SPVC from the trigeminal ganglion (TG) neurons innervating the facial skin and vomeronasal glands. Cortical input neurons exhibited clear subregional preferences. Whole-brain and spinal cord continuous imaging further demonstrated that the SPVC projects centrally via two distinct circuits that target the thalamus and the parabrachial nucleus (PB), respectively. The thalamic projections were further subdivided into dorsal and ventral circuits and exhibited clear subregional specificity within the thalamus. Additionally, the SPVC extended to the spinal cord through three major projection circuits, reaching as far as the lumbar segments. Along the whole-brain and spinal projection circuits, synaptic connections were identified in major target regions, including the submedial nucleus of the thalamus (SMT), ventral posteromedial nucleus of the thalamus (VPM), PB, and the C1 segment of the cervical spinal cord. Furthermore, tracing of SPVC pain-activated neurons revealed that they do not project to cerebellar crus 2 of the ansiform lobule (ANcr2) and paraflocculus (PFL). Conclusions High-resolution 3D imaging reveals the SPVC long-range connectome across head, brain, and spinal cord. Together, these findings establish a comprehensive connectivity framework for understanding the neural mechanisms underlying orofacial pain and provide a foundation for advances in basic sensory research and disease prevention and treatment.