The present thesis has investigated the plasticity of the sympathetic neuroeffector junction through development, and in response to environmental factors such as stressors encountered as part of experimental procedures or animal housing conditions. First, the ontogeny of sympathetic neurotransmission at a model sympathetic neuroeffector junction, the mouse vas deferens, was characterised between 14 and 42 days postnatal (PN). Excitatory junction current (EJC) amplitude increased between 28 days PN (modal EJC amplitude was 33 μV) and 42 days PN (modal EJC amplitude was 54 μV). Fewer failures were recorded from vasa deferentia of mice aged 42 days PN (failures was 0-66%) than at 28 days PN (proportion of failures was 10-75%). There was no significant difference in the ability to record the nerve terminal impulse (NTI) between 28 and 42 days PN. P2X1 receptor mRNA amplification between 10 and 42 days PN showed a
4.7-fold increase in band intensity. These results indicate that the P2X1 receptor is the most likely responsible for the EJCs recorded at the mouse vas deferens and that there is no purinergic neurotransmission at young ages.
Next, the neurotransmitter release patterns in developed mouse vasa deferentia were examined. Sympathetic nerves releasing at least two neurotransmitters, adenosine triphosphate (ATP) and noradrenaline (NA) elicit contraction of the smooth muscle in the mouse vas deferens. Several studies have indicated the presence of regional variation in the purinergic and noradrenergic contributions to sympathetic nerve-evoked contractions in rodent vasa deferentia. The relative contribution of ATP and NA to neurotransmission and contraction at the prostatic and epididymal ends of the mouse vas deferens were examined in Chapter 3. EJC success rate (extracellular indication of ATP release) from prostatically located
varicosities was eight times greater than for epididymally located varicosities. Both regions responded similarly to focal application of NA and ATP. Furthermore, the relative density and distribution of P2X1-receptor mRNA and anti-P2X1 immunostaining did not differ between the two regions. These results suggest that most epididymally located varicosities are releasing an insufficient amount of ATP to evoke detectable EJCs.
The following three experimental chapters have focussed on the plasticity of the developed sympathetic neuroeffector junction in response to animal stressor exposure. Chapter 4 aimed to determine whether a prolonged sham injection protocol could alter the levels of sympathetic neurotransmission. Three daily sham injections, equally spaced and given for 10 days resulted in a 2.1-fold increase in EJC amplitude compared to non-injected mice. Repeated injections also increased EJC success rate,
with the mean for 6-10 day treated mice being 91 ± 1.7% (mean ± SEM) compared to 40 ± 7.2% for the control. These results indicate that the efficacy of sympathetic neurotransmission can be significantly altered by the injection procedure.
Chapter 5 determined whether common animal housing conditions such as social isolation and predator (rat) presence affect sympathetic neurotransmission. Social isolation (mice housed one-per-cage in a mouse only room) significantly increased EJC amplitude by 30% compared to group housed controls (group housed mice in a mouse only room), with EJC success rate increasing by 37%. Predator threat (group housed mice in a room containing rats) had no effect, but concurrent stressor exposure (mice housed one-per-cage in a room containing rats) increased EJC amplitude and success rate significantly compared to the other three groups. Median EJC amplitude increased by 335% in the concurrent stressor
group compared to control EJC amplitude. The results suggest that common animal housing conditions can be stressful and may affect experimental results. The effects of such stressors should be given due consideration when housing experimental animals.
This thesis has more thoroughly characterised the nature of sympathetic neurotransmission and has shown that sympathetic neurotransmitter release is susceptible to environmental factors. As such, the results have implications for the advancement of animal welfare and emphasise the often overlooked possibility that common animal housing conditions and experimental procedures may actually be stressful and influence experimental results more than is often realised or expected. With the identification of other stressful housing conditions and experimental procedures, it is hoped a standard set of stress-minimising housing guidelines will be found and implemented at animal housing facilities around the world in the
interests of animal welfare.