Metabolic syndrome, first described as Syndrome X, includes a cluster of metabolic abnormalities including visceral obesity, insulin resistance, dyslipidaemia and hypertension. The pathophysiology of metabolic syndrome is complex, however, visceral adiposity and insulin resistance are thought to play a central role and both are implicated in the development of type 2 diabetes. Both metabolic syndrome and type 2 diabetes increase the risk of cardiovascular disease, by initiating and accelerating the progression of atherosclerosis, the hallmark of cardiovascular disease. Chronic low-grade inflammation of adipose tissue and vascular endothelium is well documented in type 2 diabetes and associated cardiovascular disease. In addition, stress has also been identified as one of the factors that increases the risk of cardiovascular disease. The endocannabinoid system is a physiological system that includes cannabinoid (CB1/CB2) receptors, their endogenous ligands and the enzymes responsible for their biosynthesis and degradation. Activation of the endocannabinoid system in the central nervous system leads to increased appetite and food intake while in the periphery it causes lipogenesis. Chronic over activation of the endocannabinoid system has been seen in both obesity and type 2 diabetes. Attempts to modulate the endocannabinoid system, by using CB1 antagonist/inverse agonist, rimonabant, led to positive metabolic and cardiovascular effects, but were associated with adverse psychiatric events. Cannabidiol (CBD) and delta 9 tetrahydrocannabivarin (Δ9 THCV or THCV), are two phytocannabinoids, obtained from Cannabis Sativa L. Both show a distinct pharmacological profile, separate from rimonabant and delta 9 tetrahydrocannabinol (Δ9 THC or THC), the parent compound obtained from Cannabis Sativa L. Both have potent anti-inflammatory and anti-oxidant properties and show multiple desirable cardiovascular and metabolic effects in preclinical studies. The aim of this project was to investigate the metabolic and cardiovascular effects of these two phytocannabinoids, by employing in vivo and in vitro techniques. Chapter 2 describes in detail the clinical study in type 2 diabetic subjects, where we used CBD and THCV, alone and in combination and studied their effects on various metabolic, cardiovascular and inflammatory parameters. Chapter 3 presents acute study of the cardiovascular effects of CBD in young healthy volunteers. Chapters 4 and 5 deal with the effects of CBD and THCV on mature human adipocytes and human aortic endothelial cells respectively. Last chapter includes work done on plasma samples and homogenised femoral arteries from Zucker Diabetic Fatty rats and their lean counterparts, treated with CBD. We found that THCV improves glycaemic parameters in type 2 diabetes, while CBD affects resting cardiovascular parameters and cardiovascular response to stress. CBD affects the release of leptin and IL-6 from adipocytes, while both CBD and THCV affect the release of endothelin-1 and vascular cell adhesion molecule-1 from aortic endothelial cells. CBD also shows positive impact on cytokines in diabetic rats.

Spasticity is a common presentation in a wide variety of neurological disorders like cerebral palsy (CP), multiple sclerosis, and spinal cord injury. Management of spasticity involves multiple modalities such as physical and occupational therapy, oral medicines, Botulinum toxin injection, and orthopaedic and neurosurgical intervention. Intrathecal Baclofen (ITB) therapy is one neurosurgical intervention to control spasticity in CP patients. The ITB pump is implanted subcutaneously or sub-facially in the abdomen which delivers the baclofen directly to the intrathecal space via a catheter. As a result of by-passing the blood-brain-barrier, intrathecal administration of a hundredth part of the oral baclofen dose is sufficient to relieve spasticity and therefore preventing the peripheral side-effects seen with oral administration. Although the ITB delivery systems demonstrate significant effectiveness in improving spasticity, the ITB delivery system is associated with a high complication rate which could interfere with the desired effect of ITB therapy. Therefore, in a retrospective observational study we aimed to analyse the ITB complications in a large (n=222) consecutive series of patients. The complication rate in relation to the long period of follow-up (939 pump years),was found to be similar or less than those reported in literature. Dystonia, young age and presence of gastrostomy tube were significantly associated with infective complications. Catheter complications were influenced by presence of dystonia and the surgical technique, whether it was a subfascial or subcutaneous implantation. The pump is refilled by baclofen solution. The frequency of refilling is dependent on the daily dosing regime and the concentration of the aqueous solution of baclofen in the pump. Baclofen solutions are available as commercial and compounded products. An experimental controlled study was conducted to evaluate the safety of using higher concentrations of compounded baclofen in comparison to commercial baclofen. Baclofen concentration of the compounded solution was found to be less accurate than the commercial product although it was within an acceptable range from the expected value. The number of invisible particulates was significantly higher in the compounded solution than in commercial baclofen. However, no clinical complications were reported in the compounded or in the commercial baclofen groups. As patients receiving ITB therapy may have clinical benefits of reduction of the severity of spasticity, they could also have improvements in their functional status and quality of life. A survey study to evaluate changes in functional status and heath related quality of life, showed improvement in both aspects, moreover, the improvement in the health related quality of life was more significant than the change in functional status.

Background: Influenza-like illness (ILI) and asthma are common diagnoses in the general practice and exert considerable morbidity and mortality worldwide. One potentially important strategy to reduce this is to determine whether commonly prescribed drugs modify the occurrence and outcomes of respiratory diseases including mortality. The benzodiazepines, a class of psychoactive drugs generally used in the treatment of anxiety and sleep disorders, have recently been suggested as having detrimental effects on immune response to infection, predisposing its users to increased risk of infection and mortality. These drugs have also been implicated in the pathogenesis of asthma. This thesis therefore aimed to investigate whether benzodiazepines modify the occurrence of asthma exacerbation and subsequent mortality in patients with asthma as well as ILI-related pneumonia and ILI-related mortality in patients with ILI. Methods: The Clinical Practice Research Datalink (CPRD) was used as the data source. CPRD contains the medical records of over 13 million patients prospectively collected from over 600 general practices across the United Kingdom and has linkages to the Hospital Episode Statistics database [HES] and national death registry data (the Office of National Statistics [ONS]) which were utilised in this study. CPRD-HES linked data was used to validate diagnoses of asthma exacerbation whereas ONS mortality data validated deaths identified from CPRD. Case-control and cohort study designs were used to investigate associations between benzodiazepines and the occurrence of asthma exacerbation, ILI-related pneumonia and mortality. Results: After adjusting for a wide range of potential confounders including physical and psychiatric comorbidities, and concurrent use of other drugs, exposure to benzodiazepines was associated with statistically significant increased occurrence of asthma exacerbation, ILI-related pneumonia and mortality. These associations were observed with both short term and chronic benzodiazepines use. However, the effect of individual benzodiazepines varied across the outcomes of interest with some of the associations lacking statistical significance. For instance, current use of diazepam and temazepam but not lorazepam showed statistically significant associations with increased occurrence of asthma exacerbation. Conclusion: Overall, findings of this research signal an adverse benzodiazepine effect and hence suggest that a precautionary approach should be exercised when prescribing benzodiazepines in the interim before conclusive evidence is yielded by further research, especially in patients who may already be at increased risk of asthma exacerbation or pneumonia and mortality.

Cannabinoids cause both acute and time-dependent vasodilation/vasorelaxation in a range of vascular beds. The vascular effect of cannabinoids is dependent on the cannabinoid ligand, the species studied and the vascular bed used. To date, there have been four studies that have characterised the effects of cannabinoids in humans. Therefore, the aim of this thesis was to characterise the pharmacological effects of cannabinoids in the human mesenteric artery. Written Informed consent was granted for the use of mesenteric arteries collected from patients at the Royal Derby Hospital. Arteries were dissected from mesenteric tissue and mounted on a Mulvany-Halpern myograph. Arteries were contracted using U46619 and endothelin-1. Concentration-response curves were carried out to the phytocannabinoids THC and CBD; the endocannabinoids AEA and 2-AG; the synthetic cannabinolds CP55,940 and HU-308. The underlying mechanisms of action were assessed using receptor antagonism, enzyme inhibition, endothelium denudation and ion channel manipulation. Experiments to probe the potential for cannabinoids to cause time-dependent vasorelaxation of human mesenteric arteries were also carried out. Post-hoc analysis was conducted on all acute vasorelaxation responses to assess the potential influence of patient characteristics/disease state on cannabinoid responses. All cannabinoids tested, with the exception of HU-308, caused concentration-dependent vasorelaxation of human mesenteric arteries. The synthetic cannabinoid CP55,940 had the greatest Rmax of all the cannabinoids tested. 2-AG had the greatest Rmax of the endocannabinoids tested and CBD had the greatest Rmax of the phytocannabinoids tested. Compared to animal models, cannabinoid efficacy was reduced in human mesenteric arteries. The vasorelaxant effects of 2-AG were mediated through COX-1 metabolism, prostanoid receptor activation (EP4 and IP) and ion channel modulation. The mechanisms underpinning CBD-induced vasorelaxation were CBl and TRPV1 receptor activation, NO release, the endothelium and ion channel modulation. Vasorelaxant responses to AEA were inhibited by antagonism of the CB1 receptor and a putative cannabinoid receptor located on the endothelium (CBe), nitric oxide synthase inhibition and endothelium denudation. Cannabinoid responses were reduced in patients with cardiovascular diseases/disease risk factors including ischaemic heart disease, type-2 diabetes and hypercholesterolemia. Endocannabinoid responses were reduced in patients taking NSAID medication, with some reductions in responses seen to other medication including statins and beta-blockers. CBD and AEA were tested for time-dependent vasorelaxation. Both CBD and AEA were able to cause vasorelaxatlon that gradually increased over time, this was not mediated by the PPARy receptor. This thesis concludes that cannabinoids are able to modulate vascular tone in isolated human mesenteric arteries, and this may be blunted in patients with cardiovascular disease. Furthermore, this thesis presents data suggesting that differences exist between human and animal arterial responses to cannabinoids.

Valproic acid (VPA) is extensively used for the treatment of epilepsy. This drug also functions as an inhibitor of histone deacetylase enzymes and causes the expression of growth arrest genes. It has been reported that mild to moderate cognitive impairments occur in adult patients taking VPA. This investigation aimed to examine the relationship between cognition and changes in cell proliferation within the rat hippocampus, a brain region where continued formation of new neurons is associated with learning and memory. Also, the antiproliferative function of VPA was investigated in rat and mouse hippocampal neural stem cells (NSCs) and cancer cell lines in vitro. The cognitive and antiproliferative effects of VP A were determined in adult Hooded Lister rats treated with VP A (300mg/kg) by intraperitoneal injection twice daily for 10 days. Cognition was assessed by the Novel Object Location (NOL) and contextual fear conditioning tests. Cell proliferation within the subgranular zone (SGZ) of the dentate gyrus was determined by immunostaining for Ki67. Additionally, levels of the brain-derived neurotrophic factor (BDNF), doublecortin (DCX) and the receptor Notch1 expression were measured by Western blotting. The results showed that animals treated with VPA had a hippocampal specific cognitive impairment as shown by NOL, test but not contextual fear conditioning. This was linked to a significant reduction in cell proliferation within the SGZ. Moreover, VPA treatment statistically significantly decreased levels of BDNF and Notch1, but not DCX within the hippocampus. The antiproliferative effect of VPA treatment (0.3 and 1 mM) for 24 hours was investigated in hippocampal neural stem cells (NSCs) in vitro. The numbers of neurosphere and cell proliferation assessed in VPA-treated rat and mouse hippocampal NSCs showed significantly decreased in a concentration-dependent manner. Levels of Notch1 , Sox2, nestin and c-Myc gene expression were quantified using qPCR. This revealed that 1 mM VPA reduced expression of Notch1 , Sox2 and nestin but not c-Myc. However, there were no changes in levels of these gene expressions in 0.3 mM VPA. The influence of VPA on cancer cell proliferation was examined in human Epn1, Med1 and SHSY5Y cell lines by treating with 1,2 and 3 mM VPA for 72 hours. The data of cell proliferation showed that VPA produced a significant reduction of tumour cell growth in a dose-dependent manner in all three cell lines. VPA (1, 2 and 3 mM) induced levels of Notch1 expression in SHSY5Y cell lines. Additionally, the number of dividing cells of Epn1 and Med1 treated with 2.5 mM VPA was significantly decreased compared to untreated cells using Ki67 immunostaining. However, Notch1 expression was not detected in either Epn1 or Med1 cell lines. These results indicate that VPA treatment induced cognitive deficits in rats and that this was associated with a reduction in hippocampal NSCs both in vivo and in vitro. In addition, VPA reduced BDNF and Notch1 expression in the hippocampus. Moreover, VPA induced a decrease of levels of Notch1, Sox2 and nestin gene expression in hippocampal NSCs. The present study also reveals that antiproliferative effect of VPA was associated with an increase of Notch1 expression in SHSY5Y. However, this action of VPA appeared to have no link with Notch1 expression in Epn1 and Med1 cell lines.

The current "smoking epidemic" is a global problem for governments and organisations concerned with public health. Recently, this problem has been conceptualised as one of regulation. Within the tobacco control community, there has been growing concern that the division of regulatory responsibility for conventional tobacco products (i.e. cigarettes, cigars, pipe tobacco) and alternative modes of nicotine delivery (nicotine replacement therapy products such as gums, patches and inhalers) is having adverse effects on the innovation of new medicinal products and on providing smokers with acceptable alternatives to cigarettes, the most harmful and widely used nicotine product. The 'alternatives' are mainly regulated as pharmaceuticals; therefore, must reach safety standards comparable with those required for medications rather than being compared with the known harm caused by tobacco smoking. Whilst a number of commentary and position pieces have discussed this problem, there has been little empirical work on how the current UK regulatory set-up evolved and what impacts it has. This research gap is addressed using semi-structured interviews and documentary analysis to analyse empirically the evolution and implications of divided regulatory responsibility for nicotine products in England. Adopting an actor-network theory approach, I investigate the actor-networks assembled around key non-human actors - tobacco and nicotine - focussing on developments from the 1970s until the present. In particular, I investigate the relationship between the regulatory regime and harm reduction ideas, and how they impact on the development of new medicinal nicotine products within the pharmaceutical industry. I underline the way that the regulatory regimes both shape and are shaped by the heterogeneous networks in which they are enmeshed. The thesis concludes by considering whether there are alternative approaches to regulation that would be more efficient and effective. I suggest that the regulation of recreational drug use is underpinned by 'deep conflicts in values' (Prosser 2006) and propose that further debate over the aims and limits of nicotine regulation is needed. The thesis deals with developments up until and including the 30th of November 2010.

There is considerable interest in the ability of plant-derived antioxidants to protect against oxidative damage associated with disease or exposure to toxic agents. In this study, the cytoprotection effect of the direct antioxidants quercetin (Q) and epigallocatechin-3-gallate (EGCG) and the indirect antioxidants, sulforaphane (SFN) and indole-3-carbinol (I3C) was assessed in a cellular protection assay. This assay involved two cytoprotection patterns: (a) 20-hour exposure to phytochemical followed by 5-hour exposure to t-BHP; (b) simultaneous exposure to phytochemicals and t-BHP for 5 hours. HepG2 cells were cultured to a confluent monolayer and exposed to phytochemical +/- t-BHP in serum-containing or serum-free medium, after which cell damage mediated by oxidant stress was assessed by neutral red uptake. Results showed that Q, EGCG and I3C were effective while SF was inactive and toxic to the cells by itself at high concentration during long incubation. On the other hand, short time of incubation with Q, EGCG and SF displayed identical results to prolonged exposure. However, I3C was devoid of protection activity. Moreover, results showed that serum has a major impact on antioxidant activity. The toxicity effects of t-BHP on HepG2 cells in the presence and/or absence of phytochemicals was evaluated. This assay included exposure of HepG2 cells to phytochemicals for 20 hours prior exposure to different concentrations of t-BHP for 5 hours. Results indicated that full cytoprotection was provided by Q and EGCG while partial protection was displayed by I3C and SFN. On the other hand, results showed complete cell death to have occurred at 0.4 and 0.8 mM t-BHP after 5 hours of incubation with oxidant/ in the absence of phytochemicals. Additionally, presence of serum reduces the toxic effect of t-BHP effects. The possible mechanism of the toxicity of t-BHP on HepG2 was studied to ascertain if t-BHP initiated apoptosis in HepG2 cells, through determination of activated caspase by different techniques. Basically, HepG2 cells were exposed to +/- 0.4 and 0.8 mM t-BHP for specific times. Results showed no strong evidence for apoptosis although caspase-3 activity increased significantly (p≤0.05) in treated HpG2 cells with 0.8 mM t-BHP at 150 minutes. Similarly, significant increases in ROS and lipid peroxidation in treated HepG2 cells with 0.8 mM t-BHP (p≤0.05 and 0.01 respectively) at 150 minutes were obtained. Moreover, a (non-significant) decline in GSH amount was reported. Treatment of the cells with Q and I3C under the conditions used in the cytoprotection study prevented the weak activation of caspase-3 identified by western blot. The possible mechanism (s) of protective properties of the phytochemicals in induction of phase ll detoxifying enzymes was studied by use of an intact-cell assay of quinone reductase based upon duroquinone-mediated reduction of ferricyanide. However, studies demonstrated that Q and EGCG themselves acted as intermediate electron donor in the assay, and so interfered with the assay; I3C and SFN did not share this property. In conclusion, the work presented in the thesis has indicated that the four selected compounds possess cytoprotection effect against oxidative stress induced by t-BHP under particular conditions, and has provided further insights into mechanism of toxicity of t-BHP.

The recently discovered 5-HT6 receptor has generated interest due to increasing evidence for its role in feeding, obesity, anxiety, depression and cognition. Initial studies utilising selective 5-HT6 receptor antagonists found pro-cognitive effects in various cognitive paradigms. In the last three years selective 5-HT6 receptor agonists have been developed and initial reports suggested they impaired cognition as predicted, but more recent reports have found paradoxical pro-cognitive effects in learning and memory tasks. The main aim of the current thesis was to determine the role of the 5-HT6 receptor in a conditioned emotion response (CER) task in rats. Both the effects of 5-HT6 receptor antagonists and agonists given alone, and their abilities to reverse a cholinergic- or glutamatergic-induced memory impairment were analysed. Secondly, to analyse the intracellular mechanisms involved in the behavioural effects exerted following treatment with 5-HT6 receptor ligands by examining changes in hippocampal protein expression. Pre-treatment with either the muscarinic receptor antagonist, scopolamine, or the NMDA receptor antagonist, MK-801, induced memory impairment in the 24 hour retention trial. Post-training administration of 5-HT6 receptor antagonist, SB-271046, and agonists, EMD 386088 and E-6801, had little effect on CER-induced behaviour when given alone, but both reversed the cholinergic- and glutamatergic-induced deficits. Western blot analysis revealed no significant difference between hippocampal BDNF and 5-HT6 receptor protein levels following any drug or shock treatment, but some interesting trends were observed. CER slightly increased BDNF expression, this was reduced by scopolamine and MK-801 which in turn was reversed with SB-271046 and EMD 386088. CER decreased 5-HT6 receptor expression, scopolamine caused further reduction, SB-271046 and EMD 386088 increased the expression following scopolamine. MK-801 increased 5-HT6 receptor expression, whilst SB-271046 further enhanced this expression, EMD 386088 reduced it. No significant results were observed in the proteomic studies. These findings provide further evidence for the exciting potential therapeutic use of 5-HT6 receptor compounds in the treatment of cognitive dysfunction.

Nurse independent and supplementary prescribers have legal authority to prescribe all licensed and unlicensed medicines with some minor restriction to prescribing controlled drugs. These prescribing rights are similar to those of doctors. To be effective, the integration of nurse prescribing must be consistent with the legal framework for nurse prescribing and, be acceptable to the nurse, employer, patient and healthcare team. There is little known about how prescribing is integrated in practice but agreements are potentially important to the organisation of professional work and may ultimately affect patient safety. These case studies set out to investigate how nurse prescribers integrate prescribing in primary and secondary care. Each case, a nurse prescriber,had completed the independent and supplementary prescribing course at one university between September 2004 and January 2007. Of the 26 cases recruited 13 had been qualified to prescribe for between 7 and 13 months, and 13 for 14 and 26 months. Data collected through semistructured interviews, field notes and attribute data was drawn together in case summaries. Data analysis showed effective integration to be dependent upon professional relationships and prescribing role agreements. Prescribers outlined three approaches to integrate prescribing. These were; prescribing as the opportunity presents, prescribing for specific conditions and prescribing for individuals. Prescribing as the opportunity presents reflects medical models of prescribing. Condition specific and individual approaches restrict prescribing to specific medical condition(s)or individual patients. These nurse prescribers preferred to use Independent prescribing. Reflecting this, prescribers showed higher levels of dependence on doctors than previously reported. This was most common in the first year of prescribing. Relationships between nurse prescribers and the team were important. New nurse prescribers raised unexpected issues in some intraprofessional relationships. However, it was the inter-professional relationship between nurse and doctor that determined integration. The nurse must believe, trust exists and is reciprocal to integrate prescribing in practice. Where there was an absence of trust or a concern of mistrust the nurse would not integrate prescribing.

The aryl hydrocarbon receptor is a ligand-dependent transcription factor that induces expression of a number of genes encoding drug metabolizing enzymes, such as CYP1A1, CYP1A2 and CYP1B1. Recently, it was suggested that the AhR signaling pathway may be involved in mediating the anticancer activity of novel 2-(4-aminophenyl) benzothiazole drugs in MCF-7 breast cancer cells. There is no direct proof of direct binding between these drugs and AhR, and it is also unclear how AhR signaling per se plays a role in the activity of these drugs. This study investigates the role of AhR in the mechanism of action of the benzothiazole drugs by determining the ability of these drugs to bind to the rat hepatic AhR, to induce CYP1A1 mRNA and to inhibit cell growth in rat hepatoma H4-II-E cells. The apparent binding kinetics of [3H]-TCDD to AhR in rat liver cytosol were, KD= 0.37nM and Bmax = 40 fmol/mg cytosolic protein. Using the standard assay conditions, 18 compounds competitively displaced [3H]-TCDD from specific sites, and are ligands for AhR. Induction of CYP1A1 mRNA by 5 compounds was determined in H4-II-E cells. The highest affinity ligand, IH445, was the most potent with an EC50 ~ 80-fold lower than that of TCDD (60 pM) with no detectable antagonistic activity in H4-II-E cells. The other high-affinity benzothiazoles tested were (30-100) x 103-fold less potent for inducing CYP1A1 mRNA than TCDD. The binding affinities of these compounds were 200-1000-fold higher than induction potency. For example, 5F 203 has a Ki value of 2.8 nM, induced CYP1A1 mRNA to similar maximal levels as seen with TCDD, and has an EC50 of 3 μM. The 1000-fold difference for 5F 203 between binding and CYP1A1 RNA induction was suggested to be a result of metabolism or that 5F 203 exhibits partial AhR antagonist activity. The time course effect on the CYP1A1/β-actin mRNA ratios by 5F 203 revealed that the response was increasing linearly in response to 5F 203 at 4 h treatment, indicating that the former possibilty is less likely to be a major factor. To address the second possibility, the antagonistic activity of 5F 203 on TCDD-induced CYP1A1 mRNA was investigated. H4-II-E cells were treated with increasing concentrations of TCDD ± 1μM 5F 203. The results demonstrated that 5F 203 shifted the EC50 of TCDD 100-fold to the right. Schild analysis on the antagonism of TCDD-induced CYP1A1 mRNA by different concentrations of 5F 203 provided a quantitative explanation for the 1000-fold difference between binding and induction for 5F 203. In contrast, the EC50 of 5F 203 in human MCF-7 cells was 2 nM, which is ~ 10-fold less potent than TCDD. Moreover, 5F 203 had no detectable antagonistic activity on TCDDinduced CYP1A1 mRNA. When 5F 203 was assessed for cell growth inhibition by MTT assay, it was found active in MCF-7 cells with a GI50 of 18 nM, but failed to elicit the same effect in H4-II-E cells. These results prove that 5F 203 is a potent agonist in MCF-7 cells, but a partial agonist in H4-II-E cells. The partial agonism observed with 5F 203 is a compound-specific property given that another analogue, IH 445, was found potent inducer of CYP1A1 mRNA with no antagonistic activity. The results of this study reveal species-specific partial agonism of the AhR. The potency of the cytostatic effect of 5F 203 parallel potency for inducing CYP1A1 mRNA in both cells. Moreover, both, the cytostatic effect of 5F 203 and partial agonism of AhR for inducing CYP1A1 mRNA is species-specific. Whether agonism/antagonism for the induction of CYP1A1 mRNA is related to the anticancer activity of 5F 203 remains to be elucidated.