Abstract
The adrenal medulla assists in coordinating the stress response through various neuronal, endocrine and paracrine signals. Evidence indicates that the adrenal medulla may also be responsive to signals originating from the activated immune system, such as interleukin-6, interleukin-1 and tumour necrosis factor-α, thus forming part of a bi-directional relationship. Previous research in our laboratory has indicated bovine adrenal medullary chromaffin cells are directly responsive to interleukin-6, leading to the activation of the signal transduction and activator of transcription 3 and extracellular regulated kinase pathways. Additionally, recent microarray data has indicated interleukin-6 induces alterations in a number of genes encoding neuropeptides. This thesis aims to investigate interleukin-6 induced neuropeptide gene expression by validating the microarray findings through the use of quantitative PCR, and additionally, to explore signalling mechanisms potentially involved. Primary chromaffin cell cultures were incubated for 24h in the presence or absence of interleukin-6. The total RNA was extracted and processed for qRT-PCR for selected neuropeptides (vasoactive intestinal polypeptide, galanin, stanniocalcin1, angiopoetin2, parathyroid-related peptide and gastrin releasing peptide). Immunocytochemistry was conducted to investigate expression of the latter two, both before or after treatment with interleukin-6. Cells were also treated with protein kinase inhibitors to gain an understanding of signalling pathways involved in mediating these effects. Finally, the cells were incubated with a combination of interleukin-6 and pituitary adenylyl cyclase activating polypeptide to investigate interactions. The increases in mRNA seen with the microarray data was confirmed for vasoactive intestinal polypeptide, galanin, parathyroid-related peptide and gastrin releasing peptide through qRT-PCR, and the latter two were localised to chromaffin cells by immunocytochemistry. These results confirm that interleukin-6 acts directly on chromaffin cells to increase the expression of various neuropeptides, of which two have been shown by immunocytochemistry to be present in chromaffin cells. However, it remains uncertain whether protein levels increase with interleukin-6 treatment. As some of these neuropeptides have been previously described to have anti-inflammatory actions via the adrenal cortex, this work supports existence of a stress-immune interaction. The adrenal medulla is thus implicated in preventing an exacerbated inflammatory response