Cardiac sympathetic neural signalling in hypertension

The work presented in this thesis examines the role of cardiac sympathetic signalling in the development and maintenance of hypertension in the spontaneously hypertensive rat (SHR); and the role the norepinephrine re-uptake transporter (NET) plays in the sympathetic phenotype during hypertension.
Chapter One: reviews (i) cardiac impairment observed in hypertension, (ii) the mechanisms underlying sympathetic neurotransmission and signal termination, (iii) the structure, function, mechanism of action, and role of the norepinephrine (NE) re-uptake transporter in a number of diseases related to sympathetic dysfunction including hypertension.
Chapter Two: comprises a detailed rational for the approach taken to; (i) record NET rate in single isolated cells from major sympathetic ganglia using a novel fluorescent assay (NTUA); (ii) measure norepinephrine release from isolated sympathetic cells using a method of carbon fibre based amperometry; (iii) measure autonomic function on living tissue in-vivo and in-vitro using organ bath and whole animal preparations; (iv) quantification of protein expression and protein levels within single cells and isolated ganglia.
Chapter Three: examined the cardiac autonomic phenotype of the young pre-hypertensive SHR. Baseline haemodynamics were comparable between the SHR and age matched WKY controls, apart from in-vivo heart rate which the SHR displayed a small, but significant tachycardia. Release of sympathetic neurotransmitter NE, and the co-transmitter NPY were also elevated in the young SHR compared to WKY even though the SHR showed no elevation in arterial blood pressure at this age. Tyrosine hydroxylase levels were unaltered. 3H-NE release studies also revealed there may be a dysfunction in NET in the SHR at this age, as DMI treatment normalised the difference in 3H-NE overflow between the SHR and WKY.
Chapter Four: described an investigation in to the activity of the NE re-uptake transporter between older, SHR with established hypertension and age matched normotensive WKY. A novel fluorescent assay of the monoamine transporter family was used to monitor NET activity within isolated sympathetic cells from three ganglia associated with hypertension. Direct evidence for the first time is reported that NET activity was lower in cardiac stellate neurons of the SHR compared to the WKY, but not in the other ganglia tested (superior cervical ganglion (SCG) – vascular innervation of head and neck, and celiac/superior mesenteric ganglia (CG/SMG)– renal and abdominal organs). The data support the notion that NET is regulated in a site-specific fashion, which may account for the sympathetic heterogeneity of NE spillover reported in humans.
Chapter Five: examined differences in NET activity and expression levels between four-week-old pre-hypertensive SHR and age matched WKY. The NTUA assay revealed a significantly lower rate of NET activity within stellate neurons of the young SHR compared to WKY, with no difference in NET protein expression. No difference again was seen in the other ganglia. The SCG was the only ganglia in which NET significantly decreased with age, indicating the NET dysregulation within the vasculature may develop at a later stage of hypertension than observed in cardiac ganglia.
Chapter Six: examined the ability to modulate NET activity in WKY and SHR stellate neurons in response to a number of pharmacological agents that stimulate cGMP (BNP, SNP, 8-Br-cGMP) and inhibition of PKC. These were agents previously implicated in NET modulation in other cell types. All activators of cGMP reduced NET rate to a similar degree in the WKY. No change in NET rate was observed in the SHR, indicating that NET modulation either intrinsically, and/or receptor membrane insertion, was faulty in the SHR. PKC inhibition increased NET rate in both groups.
Chapter Seven: is a concluding discussion summarising the main findings from this thesis, placing them in to physiological context, and discusses avenues of further research.

Keywords: Hypertension
Autonomic Neuroscience
Advisor: Paterson, D
Terrar, D