Vasopressin gene products are colocalised with corticotrophin-releasing factor within neurosecretory vesicles in the external zone of the median eminence of the Japanese macaque monkey (Macaca fuscata).

Akito Otubo,Yasumasa Ueda,Hirotaka Sakamoto,Sho Maejima,John F Morris,Tatsuya Sakamoto,Natsuko Kawakami

doi:10.1111/jne.12875

Abstract

Arginine vasopressin (AVP), when released into portal capillaries with corticotrophin-releasing factor (CRF) from terminals of parvocellular neurones of the hypothalamic paraventricular nucleus (PVH), facilitates the secretion of adrenocorticotrophic hormone (ACTH) in stressed rodents. The AVP gene encodes a propeptide precursor containing AVP, AVP-associated neurophysin II (NPII), and a glycopeptide copeptin, although it is currently unclear whether copeptin is always cleaved from the neurophysin and whether the NPII and/or copeptin have any functional role in the pituitary. Furthermore, for primates, it is unknown whether CRF, AVP, NPII and copeptin are all colocalised in neurosecretory vesicles in the terminal region of the paraventricular CRF neurone axons. Therefore, we investigated, by fluorescence and immunogold immunocytochemistry, the cellular and subcellular relationships of these peptides in the CRF- and AVP-producing cells in unstressed Japanese macaque monkeys (Macaca fuscata). Reverse transcription-polymerase chain reaction analysis showed the expression of both CRF and AVP mRNAs in the monkey PVH. As expected, in the magnocellular neurones of the PVH and supraoptic nucleus, essentially no CRF immunoreactivity could be detected in NPII-immunoreactive (AVP-producing) neurones. Immunofluorescence showed that, in the parvocellular part of the PVH, NPII was detectable in a subpopulation (approximately 39%) of the numerous CRF-immunoreactive neuronal perikarya, whereas, in the outer median eminence, NPII was more prominent (approximately 52%) in the CRF varicosities. Triple immunoelectron microscopy in the median eminence demonstrated the presence of both NPII and copeptin immunoreactivity in dense-cored vesicles of CRF-containing axons. The results are consistent with an idea that the AVP propeptide is processed and NPII and copeptin are colocalised in hypothalamic-pituitary CRF axons in the median eminence of a primate. The CRF, AVP and copeptin are all co-packaged in neurosecretory vesicles in monkeys and are thus likely to be co-released into the portal capillary blood to amplify ACTH release from the primate anterior pituitary.

Highlights

The present study provides the first evidence in the hypothalamus of unstressed Japanese macaque monkeys (Macaca fuscata) that corticotrophinreleasing factor (CRF), Arginine vasopressin (AVP), and the AVP-associated neurophysin II (NPII) and copeptin are all co-packaged within the neurosecretory vesicles (NSVs), and that the AVP precursor protein undergoes proteolysis during the passage of NSVs from the CRH perikarya to the median eminence (ME)
Earlier immunocytochemical studies have reported that CRF and AVP can be localised to the same parvocellular neurones in the rat[20] and mouse,[23] as well as in the human paraventricular nucleus of the macaque hypothalamus (PVH).[12,21,35]
It appears likely that CRF and the other AVP precursor components (NPII and copeptin) are copackaged within the same NSV in the primate hypothalamus, there has far been no anatomical evidence

Summary

| MATERIALS AND METHODS

The hypothalamic neuroendocrine response to stress is an important survival mechanism in vertebrates. Any clear evidence for the function of the secreted copeptin is currently lacking.[19] The present study provides the first evidence in the hypothalamus of unstressed Japanese macaque monkeys (Macaca fuscata) that CRF, AVP, and the AVP-associated NPII and copeptin are all co-packaged within the NSVs, and that the AVP precursor protein undergoes proteolysis during the passage of NSVs from the CRH perikarya to the ME. Monkeys were maintained under a 12:12 hour light/dark cycle (lights off 8.00 pm) in a temperature-controlled (22-24oC) room. These animals were checked and shown to be free of specific pathogens. All efforts were made to minimise animal suffering and reduce the number of animals used in the study

| Experimental procedures

Findings

| DISCUSSION