The experiments described in this thesis have investigated the role of nerve growth factor (NGF) acting on spared nerve fibres in the pathophysiology of neuropathic pain, caused by a partial nerve injury of the mental nerve, a branch of the trigeminal nerve that innervates the skin and mucosa of the lower lip. The experiments have tested the hypothesis that an over-supply of NGF acts on a reduced population of mental nerve fibres and causes phenotypic changes in the population that could underlie symptoms of neuropathic pain. A behavioural testing protocol was developed to investigate the presence and severity of neuropathic pain to thermal and mechanical stimuli after a chronic constriction injury of the mental nerve. The concentration of NGF in the skin of the lower lip was investigated with an ELISA (enzyme-linked immunosorbant assay) and the neurochemistry of the remaining sensory nerve fibres that innervate the skin of the lip was investigated quantitatively using triple-labelling immunofluorescence. Experiments to determine the time-course of the development of neuropathic pain were conducted initially, and when symptoms were maximal (11, 14 and 21 days post-operatively) further studies on NGF concentration and neurochemical phenotype were performed. A chronic constriction injury of the mental nerve produced cold hyperalgesia in the ipsilateral lower lip 1I days post-injury, although at this time the NGF concentration in the lower lip did not differ between ipsilateral and contralateral sides (ipsilateral = 45±6(SEM) pg/mg, contralateral = 47±7 pg/mg). NGF concentration was significantly increased ipsilaterally compared to contra laterally 14 days post-operatively (ipsilateral = 111±23 pg/mg, contralateral = 69± 13 pg/mg), however, at this time-point there was no behavioural evidence of neuropathic pain. By 21 days post-injury, NGF concentration both ipsilaterally and contralaterally was elevated compared to data for 11 days but there was no significant difference between sides (ipsilateral = 181±31 pg/rng, contralateral = 205±30 pg/rng) and there was also no behavioural evidence of neuropathic pain. Quantitative immunohistochemistry of samples 21 days post-injury showed a significant increase on the ipsilateral side (ipsilateral = 85± 17%, contralateral = 46±5%) in the proportion of nerve fibres that were double-labelled for trkA (the high-affinity NGF receptor) and Calcitonin gene-related peptide (CGRP) and this change was absent at 11 and 14 days post-injury. The data from these investigations suggest that cold hyperalgesia following partial injury to the mental nerve is not induced from an altered concentration of NGF in the skin and that a different mechanism may cause this symptom. Transiently increased NGF on the ipsilateral side may have caused an increased proportion of CGRP immunoreactive fibres in the trkA population and this difference in innervation pattern coincided temporally with mechanical nociceptive thresholds on the ipsilateral side that were lower than those on the contralateral side (due to a lack of habituation which occurred on the contralateral side of chronic constriction injuries and on both sides of sham operated animals).

Diabetic patients present with initial hypersensitivity within peripheral located skin, followed later by a switch to nocicpetive insensitivity. Loss of nociception coupled with reduced neurovascular responses to injury can result in ulceration, particularly of the foot. One of the neurovascular responses to injury is axon reflex vasodilation of the affected skin, mediated by calcitonin gene-related peptide (CGRP) release from the afferent nerve. A working protocol was established to measure neurogenic vasodilation evoked by short electrical stimulation of the saphenous nerve. In the streptozotocin-induced diabetic rat model, four weeks of untreated diabetes resulted in an approximate 40% reduction in the skin blood flow response, with a further 45% reduction by eight weeks. Vasodilation to acute administration of CGRP was unchanged at four weeks and reduced by approximately 60% by eight weeks. Antioxidant (alpha-lipoic acid) or ACE-inhibitor (lisinopril) treatment prevented the progression of deterioration between four and eight weeks of diabetes, but could not prevent the development of the neurogenic deficit over the first four weeks of the disease. IGF-I replacement therapy not only prevented the deterioration of the neurogenic response between four and eight weeks of diabetes, but it completely normalised the functional deficit to control levels. Diabetic rats also demonstrated tactile allodynia, thermal hyperalgesia and reduced CGRP content of dorsal root ganglion cells, all of which were normalised by IGF-I treatment. The data indicate that an improvement in oxidant status and blood flow may be effective in halting the early progression of diabetic neuropathy and that IGF-I replacement therapy may prove to be efficacious in the alleviation of painful neuropathy and functional hyperaemia in the diabetic foot.

Congenital Hypomyelination (CH) is the most severe demyelinating form of Hereditary Motor and Sensory Neuropathies and manifests at birth in human. Some subtypes of CH are due to dommant mutations in the gene coding for PO glycoprotein, which fiinctions as a homophilic adhesion protein, responsible for compaction of opposing myelin lamellae. By homologous recombination in ES cells, we have generated a mouse containing a nonsense mutation m the intracellular portion of PO (Q215X) that, in the heterozygous state, is associated with CH neuropathy in humans.

Aims: Identification of neurophysiological or skin innervation biomarkers which can be used to assess and monitor progression of diabetic sensory polyneuropathy (DPN) and chemotherapy-induced neuropathy (CIPN). Sensitive and robust measures are needed to detect changes in the relatively short duration of clinical trials aimed to modify progression of neuropathy. Methods: 40 patients with DPN were studied longitudinally over 1 year, and 33 patients with CIPN in a cross-sectional study. Clinical assessments, questionnaires, quantitative sensory testing, histamine-induced skin flare, nerve conduction studies and contact heat evoked potentials were measured. Repeat skin biopsies were performed at a 6 month interval to quantify intra- (IENF) and sub-epidermal (SENF) nerve fibres immunoreactive for PGP 9.5 (pan-neuronal marker), TRPV1 (heat and capsaicin receptor) and GAP-43 (marker of regenerating fibres) in the DPN group, and at baseline in the CIPN group. Results: There was no change in symptoms and sensory tests in the DPN group. However, there was a significant reduction in IENF and SENF for both PGP 9.5 and TRPV1 fibres in the second DPN skin biopsy (n = 29 had repeat biopsy). GAP- 43 fibres were present in the dermis and remained unchanged. Patients in the CIPN group had less painful neuropathy, but similar abnormalities on examination and sensory tests. Despite this, a preserved number of IENF and SENF were seen in the CIPN group, with abnormal morphology. This has not been reported previously. Conclusion: PGP 9.5 and TRPV1-immunoreactive nerve fibres in sequential skin biopsies provide objective markers of progression of neuropathy, while the preserved GAP 43-immunoreactive fibres may detect enhanced regeneration. Novel findings in the CIPN group suggest prevention of degeneration and restoration of function should be the treatment strategy, rather than enhancing regeneration.

No description available.

Guillian-Barré Syndrome (GBS) is the world’s leading cause of neuromuscular paralysis occurring in serologically and pathogenically distinct forms. GBS is believed to have an autoimmune basis, where antibodies raised during antecedent infections (eg Campylobacter jejuni) cross-react with self antigens, exemplifying the process of molecular mimicry. These self-antigens are gangliosides, which are glycolipid structures enriched in peripheral nerve in specific membrane compartments termed lipid rafts. To date, successful murine models of anti-GD1a and anti-Gq1b ganglioside mediated neuropathy exist. Clinical evidence supports the involvement of anti-GM1 antibodies in nerve injury, however generation of anti-GM1 antibody mediated neuropathy models remain an enigma, and to date, the only successful model is based in Japanese rabbits. This thesis aims to address the controversies surrounding anti-GM1 antibody mediated neuropathy by utilising a panel of anti-GM1 antibodies of differing specificity, and explores how the stereometric interactions of GM1 with lipid raft species underpin the pathogenic potential of these antibodies.

- / Our purpose was to assess the function of fast somatic nerve fibres, the sympathetic sudomotor system and the function of parasympathetic heart con-trol in patients with partial or generalized seizures, receiving various therapeu-tic regimes, by means of clinical routine neurophysiology. We studied 73 pati-ents (mean age 37,7±12,9 years) and 73 matched healthy controls by clinical and electrophysiological examinations. To assess the function of fast somatic nerve fibres we studied the motor conduction of the peroneal nerve (distal latency, amplitude of action potential, conduction velocity) and the sensory conduction of the ulnar and sural nerves (amplitude of action potential, conduction velocity) in all patients and controls. To assess the sympathetic sudomotor system we studied the sympathetic skin response (SSR) elicited in the hand and foot by electrical and magnetic stimulation. The electrical stimulation applied in all patients and controls but the magnetic stimulation applied in 61 patients and 72 contols (exepted pati-ents and controls wich had something metallic in their body such as surgical clips, pace-maker, metallic valve). To assess the function of parasympathetic heart contol all the patients and the controls were subjected to a battery of three cardiovascular tests: a) heart rate variability during six deep breaths/min [algorithm:(max-min)R-R Interval x 100/mean R-R Interval] b) Valsava maneuver (ratio of the shortest RR interval during phase II to the longest RR interval in phase IV) c) heart rate response to standing up(ratio 30th beat/15th beat). Compared to controls, patients showed a significant abnormality of motor (prolongation of distal latency) and sensory (slowing of ulnar conduction velo-city, reduction of amplitude of sural action potential) conduction. Reason and duration of epilepsy, type and frequency of seizyre and polytherapy didn’t ap-pear to have a negative effect on motor or sensory conduction. Particular antiepileptic drugs, oxcarbazepine, phenytoin, topiramate, lebetiracetam, ap-peared to have a negative effect on sensory conduction. Clinical sings of neu-ropathy were disclosed in 13,7% of patients, while at least one abnormal sen-sory parameter was evident in 11%. In the patient group as opposed to the control group, there was a latency prolongation of sympathetic skin response recorded in hand and foot to elec-trical and magnetic stimulation. At least one SSR abnormality was shown in 28 (38,3%) patients and 9 (12,3%) of these had absent SSR from the hand or/and the food with electrical or/and magnetic stimulation. Reason & duration of epi-lepsy, type, frequency & age at onset of seizure, time from last seizure and po-lytherapy didn’t appear to have a negative effect on sympathetic sudomotor function. Particular antiepileptic drugs, topiramate and phenytoin, appeared to have a negative effect on sympathetic sudomotor function. Finally, the patient group showed significantly difference in all parasympa-thetic parameters studied compared to controls. Overall, 42,2% of patients had abnormal measurement in at least one of these parameters and [algorithm:(max-min)R-R Interval x 100/mean R-R Interval] was the most fre-quently abnormal one (31,5% of patients). Reason & duration of epilepsy, fre-quency, type & age at onset of seizure, time from last seizure and polytherapy didn’t appear to have a negative effect. Particular antiepileptic drugs, oxcarba-zepine and phenytoin appeared to have a negative effect on fuction of para-sympathetic heart control. Overall, 50 of the 73 patients had abnormal measurement in at least one of all parameters wich we studied. Our results suggested that simple neurophysiological tests, suitable for screening purposes, were able to demomstrate abnormalities of the fast so-matic fibers in peripheral nerves, of the sympathetic sudomotor system and of the parasympathetic heart control.