Calcitonin Receptor And Receptor Activity-Modifying Protein 1 Expression In The Human Medulla

Rebekah L Bower, Henry J Valdvogel, Richard LM Faull, Debbie L Hay. University of Auckland, Auckland, New Zealand.

The calcitonin receptor (CTR) is a family B G protein-coupled receptor which responds to calcitonin. The CTR may heterodimerise with accessory proteins called receptor activity-modifying proteins (RAMPs), producing amylin receptor phenotypes with unique pharmacologies. Amylin is a peptide hormone released postprandially which is involved in glucoregulation and satiety. The actions of amylin are postulated to be centrally mediated, but the expression of CTR and RAMPs have never been studied in the human (h) brain. The medulla oblongata is an autonomic control centre which possesses regions devoid of a blood brain barrier that are thought to bind and respond to circulating hormones such as amylin (Sexton, 1994). Therefore, the aim of this study was to determine whether amylin receptor components (hCTR and hRAMP1) are expressed in the human medulla.

Antibodies for hCTR (9B4) and hRAMP1 (844) were obtained from Welcome Receptor Antibodies and Merck (kindly provided by Chris Salvatore), respectively. Six healthy human brains were obtained from the Neurological Foundation of New Zealand human brain bank with the approval of the University of Auckland Human participant’s ethics committee. Brains were fixed by perfusion through the carotid and basilar arteries with phosphate-buffered saline and 1% sodium nitrite followed by 15% formalin fixative in 0.1M phosphate buffer. Tissue blocks were dissected from the brain and post-fixed in 15% formalin fixative for 24 hours. 20% followed by 30% sucrose in 0.1M phosphate buffer with 0.1% sodium azide over 2 weeks was used to cryo-protect the tissue blocks. Tissue blocks were rapidly frozen on powdered dry ice and stored at -80°C until required for immunohistochemical processing. Antibody specificity was confirmed in cell culture prior to immunostaining and immunohistochemistry was performed on free-floating 50µm sections of the human medulla.

Expression of both hCTR and hRAMP1 overlapped in regions of the nucleus of the solitary tract, external cuneate nucleus, and the inferior olivary nucleus. Therefore, hCTR and hRAMP1 could be interacting to yield an amylin receptor in these regions. The nucleus of the solitary tract is known to signal upstream to higher order autonomic control centres such as the hypothalamus. Therefore, this region may mediate amylin’s actions of satiety. The main functions of the external cuneate and the inferior olivary nuclei are proprioception of the upper limbs and control of movement, respectively. The function of expressed hCTR and hRAMP1 in these regions is as yet unclear. hRAMP1 and not hCTR was highly expressed in the lateral reticular nucleus, known to have involvement in cardiovascular functions. hRAMP1 could also be associating with a related receptor (the calcitonin receptor like receptor) to yield calcitonin gene-related peptide (CGRP) receptors. This is the first report of CTR and RAMP1 expression in these regions of the human brain, which may help explain the actions of amylin and suggest novel functions.

Sexton, P. M. (1994). Neuroscience, 62, 553-67.