Anti-CALCA polyclonal antibody

Calcitonin gene-related peptide (CGRP alpha, encoded by Calca) is a classic marker of nociceptive dorsal root ganglia (DRG) neurons. Despite years of research, it is unclear what stimuli these neurons detect in vitro or in vivo. To facilitate functional studies of these neurons, we genetically targeted an axonal tracer (farnesylated enhanced green fluorescent protein; GFP) and a LoxP-stopped cell ablation construct (human diphtheria toxin receptor; DTR) to the Calca locus. In culture, 10-50% (depending on ligand) of all CGRP alpha-GFP-positive (+) neurons responded to capsaicin, mustard oil, menthol, acidic pH, ATP, and pruritogens (histamine and chloroquine), suggesting a role for peptidergic neurons in detecting noxious stimuli and itch. In contrast, few (2.2 +/- 1.3%) CGRP alpha-GFP(+) neurons responded to the TRPM8-selective cooling agent icilin. In adult mice, CGRP alpha-GFP(+) cell bodies were located in the DRG, spinal cord (motor neurons and dorsal horn neurons), brain and thyroid-reproducibly marking all cell types known to express Calca. Half of all CGRP alpha-GFP(+) DRG neurons expressed TRPV1, similar to 25% expressed neurofilament-200, <10% contained nonpeptidergic markers (IB4 and Prostatic acid phosphatase) and almost none (<1%) expressed TRPM8. CGRP alpha-GFP(+) neurons innervated the dorsal spinal cord and innervated cutaneous and visceral tissues. This included nerve endings in the epidermis and on guard hairs. Our study provides direct evidence that CGRP alpha(+) DRG neurons respond to agonists that evoke pain and itch and constitute a sensory circuit that is largely distinct from nonpeptidergic circuits and TRPM8(+)/cool temperature circuits. In future studies, it should be possible to conditionally ablate CGRP alpha-expressing neurons to evaluate sensory and non-sensory functions for these neurons.

It is well known that type 1 diabetes is associated with a decrease in bone mass and delayed healing of fractures in human and in animal models of type 1 diabetes. Using well- and poorly compensated diabetic BB/ O(ttawa) K(arlsburg) rats spontaneously developing insulin-dependent type 1 diabetes, it was recently shown that, in contrast to all other tissues studied, bone is most influenced by metabolic state and seems to be regulated in a manner different from other organs. Therefore, we studied the expression of additional genes (Bmp-1, Bmp- 4, Vegf, Bglap, Il-1b, Infg, Tnfa, Calca, Sp1, Yy1) in bone of nondiabetic BB rats compared with newly diagnosed and well- and poorly compensated diabetic rats as well as two nondiabetes- prone congenic BB. SHR rats, BB rat-related (WOKW) and -unrelated rat strains (F344). Six males of each group were euthanized, the tibial bone was removed, and total RNA was extracted, transcribed in complementary DNA, and used for real-time PCR. In a comparison of nondiabetic with diabetic groups, the relative gene expression was reduced by >80% in newly diagnosed and in well- compensated diabetic BB/ OK rats. The gene expression in poorly compensated rats increased significantly in 7 of 10 genes and was comparable with those of nondiabetic BB/ OK rats. In a comparison of gene expression between diabetes-prone BB/ OK and nondiabetes- prone BB. 1K, BB. 4S, WOKW, and F344 rats, there were no significant differences between newly diagnosed and well- compensated BB/ OK diabetic rats and nondiabetic BB. 1K, BB. 4S, WOKW, and F344 rats. On the basis of these findings, we concluded that spontaneous diabetes influences bone gene expression in BB/ OK rats, which may be attributed to the genetically determined autoimmune process not only affecting pancreatic beta-cells but also bone formation and resorption.