Using tissue clearing and light sheet fluorescence microscopy for the three-dimensional analysis of sensory and sympathetic nerve endings that innervate bone and dental tissue of mice.

Bone and dental tissues are richly innervated by sensory and sympathetic neurons. However, the characterization of the morphology, molecular phenotype, and distribution of nerves that innervate hard tissue has so far mostly been limited to thin histological sections. This approach does not adequately capture dispersed neuronal projections due to the loss of important structural information during three-dimensional (3D) reconstruction. In this study, we modified the immunolabeling-enabled imaging of solvent-cleared organs (iDISCO/iDISCO+) clearing protocol to image high-resolution neuronal structures in whole femurs and mandibles collected from perfused C57Bl/6 mice. Axons and their nerve terminal endings were immunolabeled with antibodies directed against protein gene product 9.5 (pan-neuronal marker), calcitonin gene-related peptide (peptidergic nociceptor marker), or tyrosine hydroxylase (sympathetic neuron marker). Volume imaging was performed using light sheet fluorescence microscopy. We report high-quality immunolabeling of the axons and nerve terminal endings for both sensory and sympathetic neurons that innervate the mouse femur and mandible. Importantly, we are able to follow their projections through full 3D volumes, highlight how extensive their distribution is, and show regional differences in innervation patterns for different parts of each bone (and surrounding tissues). Mapping the distribution of sensory and sympathetic axons, and their nerve terminal endings, in different bony compartments may be important in further elucidating their roles in health and disease.

5-HT(1D) receptor immunoreactivity in the sphenopalatine ganglion: implications for the efficacy of triptans in the treatment of autonomic signs associated with cluster headache.

OBJECTIVE: To determine if 5-HT(1D) receptors are located in the sphenopalatine ganglion. BACKGROUND: While the 5-HT(1D) receptor has been described in sensory and sympathetic ganglia in the head, it was not known whether they were also located in parasympathetic ganglia. METHODS: We used retrograde labeling combined with immunohistochemistry to examine 5-HT(1D) receptor immunoreactivity in rat sphenopalatine ganglion neurons that project to the lacrimal gland, nasal mucosa, cerebral vasculature, and trigeminal ganglion. RESULTS: We found 5-HT(1D) receptor immunoreactivity in nerve terminals around postganglionic cell bodies within the sphenopalatine ganglion. All 5-HT(1D) -immunoreactive terminals were also immunoreactive for calcitonin gene-related peptide but not vesicular acetylcholine transporter, suggesting that they were sensory and not preganglionic parasympathetic fibers. Our retrograde labeling studies showed that approximately 30% of sphenopalatine ganglion neurons innervating the lacrimal gland, 23% innervating the nasal mucosa, and 39% innervating the trigeminal ganglion were in apparent contact with 5-HT(1D) receptor containing nerve terminals. CONCLUSION: These data suggest that 5-HT(1D) receptors within primary afferent neurons that innervate the sphenopalatine ganglion are in a position to modulate the excitability of postganglionic parasympathetic neurons that innervate the lacrimal gland and nasal mucosa, as well as the trigeminal ganglion. This has implications for triptan (5-HT(1D) receptor agonist) actions on parasympathetic symptoms in cluster headache.

Peer Tutoring for Anatomy Workshops in Cambodia.

Historical loss of staff and teaching resources in Cambodia has resulted in significant challenges to anatomy education. Small group anatomy teaching opportunities are limited. A visit to Cambodia by a teaching team from the University of Melbourne in 2010 demonstrated it was possible to implement well-resourced anatomy workshops for this purpose. However, continuation of the workshop program was inhibited by the limited number of local teaching staff. In 2015, another team from the University of Melbourne returned to Cambodia to implement anatomy workshops that incorporated peer tutoring. The objective was to improve teacher-to-student ratios and to demonstrate that interactive anatomy workshops could be delivered successfully despite low staff numbers. The anatomy workshops were attended by 404 students of Medicine, Dentistry, Nursing, and Midwifery at the University of Puthisastra. Medical students were invited to act as peer tutors for nursing students. A five-point Likert scale questionnaire was used to determine student satisfaction with both the workshops and peer tutoring. The overwhelming majority were positive about the workshops and keen for them to continue. Almost all medical students who acted as peer tutors agreed or strongly agreed that this role increased their anatomical knowledge (98%) and confidence (94%). Most nursing students agreed or strongly agreed with statements that they would like peer tutoring to continue (94%) and that they would like to be peer tutors themselves (88%). This report demonstrates that peer tutoring could be an effective tool in educational settings where poor staff-to-student ratios limit delivery of interactive workshops.

Collaborative development of anatomy workshops for medical and dental students in Cambodia.

After Phnom Penh was liberated from the Khmer Rouge in 1979, health science education in Cambodia had to be completely rebuilt. In this article, the authors report the results of a teaching collaboration between the University of Melbourne (Australia), the International University (Cambodia), and the University of Health Sciences (Cambodia). The main objectives in this collaboration were to provide the opportunity for dental and medical students in Cambodia to attend resourced anatomy workshops and to provide an opportunity for anatomy teachers in Cambodia to gain experience in implementing anatomy workshops of the style that are routinely used in the medical and dental curricula at the University of Melbourne. Experienced anatomy educators from the Department of Anatomy and Cell Biology, University of Melbourne, designed and resourced a series of workshops and then delivered these in collaboration with Cambodian teaching staff in Phnom Penh. The Cambodian students who participated in the workshops were incredibly engaged and enthusiastic. The students' evaluations (by questionnaire) indicated a very positive response to the workshops. All of the workshop resources were donated to the two universities so that the staff could continue to implement similar workshops, and plans were developed to continue our collaboration by developing more resourced workshops for this purpose in the future. Two staff members from Cambodia will travel to Melbourne to participate in anatomy workshops and dissection classes at the University of Melbourne. We hope that this extension of the collaboration provides further support and impetus for the development of anatomy education in Cambodia in the future.