Behavioural Ccnsequences and neural correlates of bladder inflamation in the laboratory rat

Abstract

The treatment and management of pain continues to reflect the complex nature of pain itself. At present, pain cannot be definitively tested for, only inferred through a combination of sensory and self-­‐report measures, such as quantitative sensory testing procedures and validated questionnaires, which survey various aspects of the pain experience in more detail, as well as the impact of pain on daily life at home and in the workplace. Whilst clinical sensory testing outcomes are analogous to in vivo evoked sensory data, modelling the impact of pain on quality of life is more challenging. One approach is to use an ethological paradigm, i.e. one that measures a behaviour that reflects an innate environmental response. Tests such as the open field and burrowing measure how pain affects threat avoidance (thigmotaxis in the open field) and tunnel maintenance (burrowing) in laboratory rats. Studies have shown their efficacy in detecting subtle behavioural deficits in experimental models of pain, including response to analgesics. Using ethological outcome measures also avoids applying clinical terminology to animal behaviour, and should be used in conjunction with classical sensory reflex-­‐driven assays, such as hotplate and von Frey filaments, to yield a global picture of how experimental pain states affect wellbeing and quality of life. This study examines how acute visceral inflammation, persistent visceral inflammation, and drug-­induced neuropathy (d4T-­ antiretroviral nucleoside) alter the behaviour of adult female Wistar rats in the open field paradigm. It also investigates how the behavioural phenotype of naïve male Wistar rats in the open field paradigm. It also investigates how the behavioural phenotype of naïve male Wistar rats responds to repeated open field exposure.

Significant increases in thigmotaxis were observed following acute bladder inflammation, and repeated exposure to open field in naïve rats. There was no effect on either open field behaviour or evoked sensory measures in female rats treated with d4T, and acute bladder inflammation failed to alter burrowing behaviours. As d4T has been previously shown to alter thigmotactic behaviour, this data suggests that females show different sensitivity compared to males.

To determine whether visceral inflammation influences behaviour in a dose-­‐dependent manner, bladder tissue from the visceral inflammation groups was assayed for levels of cytokines using an RNA microarray featuring 92 inflammatory cytokines and 4 housekeeping genes. The greatest effect was detected following acute inflammation, with both models showing increased levels of CCL12, CCL7, and IL-­‐1β when compared to naive tissue. To correlate the changes in behaviour with a neural substrate, c-­‐Fos immunoreactivity in the amygdala was measured in both visceral inflammation groups. No significant activational differences were seen following acute bladder inflammation, whereas persistent visceral inflammation significantly increased c-­‐Fos immunoreactivity in the caudal regions of the capsular central amygdala. Visceral inflammation is associated with peripheral increases in inflammatory cytokines, and thigmotactic behaviour in the open field. Differences in activation in the amygdala were lower than expected due to high levels of variation associated with inflammatory stimuli at the time-­‐point tested. Significant variation was particularly seen in repeated visceral inflammation, suggestive of the biphasic behavioural response to stress e.g. attack or defend phenotypes. Further examination of these differences at the individual level could shed light on the process of pain chronification, and ultimately help understand why only some individuals develop chronic pain.