Abstract:
The calcitonin family of peptides modulate a range of physiological processes that make them
amenable to therapeutic exploitation. The neuropeptide calcitonin gene-related peptide (CGRP) has
particular value as it plays a major role in the pathophysiology of migraine. The related hormone amylin
is also therapeutically relevant for both diabetes and obesity due to its effects on satiation and
glucoregulation. However, both CGRP- and amylin-based treatments have limitations including modest
efficacy. Improving the clinical value of these two peptides is, in part, restricted by incomplete
understanding of the molecular pathways through which they mediate specific (patho)physiological
processes.
CGRP and amylin have sites of action in the brain, where radioligand binding reveals widespread
binding sites for both peptides. The molecular correlates responsible for these binding sites, which
represent cellular receptors, are not yet known. The in vitro pharmacology of CGRP and amylin
receptors is complex due to their unusual heterodimeric nature. The canonical receptor for CGRP
comprises a G protein-coupled receptor, known as the calcitonin-like receptor (CLR), coupled to a
receptor activity-modifying protein (RAMP1). Interestingly, while the related calcitonin receptor (CTR)
complexed with RAMP1 is termed an amylin 1 (AMY1) receptor, it is potently activated by both CGRP
and amylin in vitro. It is not clear if the AMY1 receptor is a genuine receptor for both peptides in vivo.
A foundational step in understanding the role of the AMY1 receptor in CGRP and amylin biology is
clearly defining the expression patterns of the two receptor components (CTR and RAMP1) in the brain,
where both peptides are active. Where it is available, expression information is often incomplete.
Therefore, the aim of this thesis was to examine the tissue localisation of CTR and RAMP1 in rat brain.
Select regions of the brain were targeted based on a comprehensive literature review including peptide
binding sites, RNA and protein expression and physiological relevance. The experimental strategy
chosen was immunohistochemistry, which uses antibodies as specific probes. Therefore, an important
first step was characterising the antibodies chosen for their selectivity and suitability, using
complementary cell-based methods. In the rat brain, robust CTR-like immunoreactivity was observed
in many regions relevant for CGRP and amylin, particularly in the brainstem. RAMP1-like
immunoreactivity was also robust but varied between antibodies. Ultimately, transgenic mouse models
revealed that RAMP1-like immunoreactivity could not be confidently attributed to the presence of
RAMP1. This has implications for previous studies and our overall understanding of RAMP1 expression
in the brain. This thesis offers an expanded perspective on the receptors for CGRP and amylin in the
brain, which is critical for advancing the understanding of these peptides’ biology as well as their clinical
utility.