Peripheral neuropathy in mice with neuronal nitric oxide synthase gene deficiency

Abstract

Abstract. Evidence for the important role of the potent oxidant peroxynitrite in peripheral diabetic neuropathy and neuropathic pain is emerging. This study evaluated the contribution of neuronal nitric oxide synthase (nNOS) to diabetes-induced nitrosative stress in peripheral nerve and dorsal root ganglia, and peripheral nerve dysfunction and degeneration. Control and nNOS -/-mice were made diabetic with streptozotocin, and maintained for 6 weeks. Peroxynitrite injury was assessed by nitrotyrosine and poly(ADP-ribose) immunoreactivities. Peripheral diabetic neuropathy was evaluated by measurements of sciatic motor and hind-limb digital sensory nerve conduction velocities, thermal algesia, tactile allodynia, and intraepidermal nerve fiber density. Control nNOS -/-mice displayed normal motor nerve conduction velocity and thermal response latency, whereas sensory nerve conduction velocity was slightly lower compared with non-diabetic wild-type mice, and tactile response threshold and intraepidermal nerve fiber density were reduced by 47 and 38%, respectively. Both diabetic wild-type and nNOS -/-mice displayed enhanced nitrosative stress in peripheral nerve. In contrast to diabetic wild-type mice, diabetic nNOS -/-mice had near normal nitrotyrosine and poly(ADP-ribose) immunofluorescence in dorsal root ganglia. Both diabetic wild-type and nNOS -/-mice developed motor and sensory nerve conduction velocity deficits and thermal hypoalgesia although nNOS gene deficiency slightly reduced severity of the three disorders. Tactile response thresholds were similarly decreased in control and diabetic nNOS -/-mice compared with non-diabetic wild-type mice. Intraepidermal nerve fiber density was lower by 27% in diabetic nNOS -/-mice compared with the corresponding nondiabetic group, and by 20% in diabetic nNOS -/-mice compared with diabetic wild-type mice. In conclusion, nNOS is required for maintaining the normal peripheral nerve function and small sensory nerve fibre innervation. nNOS gene deficiency does not protect from development of nerve conduction deficit, sensory neuropathy and intraepidermal nerve fiber loss.

Phenotypic Changes in Diabetic Neuropathy Induced by a High-Fat Diet in Diabetic C57Bl/6 Mice

“…It is unclear why the results from the current study specifically differ from many other previous reports of slowed NCV in STZ-diabetic C57Bl/6 mice of similar age and diabetes duration [ 43 , 44 , 49 , 50 ], but a major contributing factor may be the small number of animals examined, or differences in antioxidant and macronutrient composition of the standard diet, or even the mouse supplier (Charles River versus Jackson Laboratories). As mentioned above, dietary conditions may strongly impact the neuropathy phenotype.…”

“…However, the duration of high-fat feeding was longer (12 or 16 weeks) in the previous studies [ 24 , 25 , 41 ] compared to 8 weeks of high-fat feeding in the current study. Although MNCV was significantly increased in nondiabetic mice fed the high-fat diet compared to their counterparts on the standard diet, MNCV in high-fat-fed nondiabetic mice was 53.1 ± 1.9 in the current study which is similar to previous reports of MNCV (approximately 50 m/s) in nondiabetic mice fed a standard diet [ 29 , 43 , 44 , 52 ]. Although no known reports of high-fat feeding have shown increased MNCV in nondiabetic mice, it is possible that increased fat intake during a critical stage of development (8–16 weeks) may affect myelination and transiently increase MNCV at this specific stage of development (16 weeks of age).…”

“…Although sensory and motor NCV were reduced in diabetic mice fed the high-fat diet compared to their nondiabetic counterparts, diabetes alone did not significantly affect NCV in the current study. Sensory and motor nerve conduction deficits are a well-documented consequence of diabetes in rodents [ 28 , 43 , 44 , 46 – 53 ] and several studies report diabetes-induced slowed SNCV and MNCV in STZ-diabetic C57Bl/6 mice. However, reports of diabetes-induced nerve conduction deficits can be variable, ranging from no change in SNCV [ 29 , 34 , 35 ] or MNCV [ 54 , 55 ] up to approximately 30% and 20% reductions in sensory and motor NCV, respectively [ 52 ].…”

“…As mentioned above, dietary conditions may strongly impact the neuropathy phenotype. For example, previous reports of slowed NCV in STZ-diabetic C57Bl/6 mice of similar age and diabetes duration obtained mice from Jackson Laboratories or other vendors [ 43 , 44 , 49 , 50 ] while the C57Bl/6 mice in the current study were obtained from Charles River. In addition, the diet the animals were fed during the previous studies [ 43 , 44 , 49 ] was an unspecified standard mouse chow from PMI Nutrition International while the standard diet in the current study was from Harlan Teklad (8604) and were likely different in antioxidant and macronutrient composition.…”

Role of nitrosative and oxidative stress in neuropathy in patients with type 2 diabetes mellitus

“…There is no doubt that the availability of NO • is decreased in type 2 diabetic patients and the results of this study add a further finding of significant high level of peroxynitrite (ONOO • ), an end-product of non-enzymatic reaction between nitric oxide (NO • ) and superoxide anion (O2 •- ) radicals. [ 20 – 22 ] The non-significant correlation between oxidative or nitrosative stress with nerve conduction velocity could be attributed to the wide variation in duration of illness, small sample size of patient, and the levels of lipoprotein oxides and ONOO • in biological fluids rather than in nerve tissue. In the non-diabetic experimental animal model the significant increase lipid peroxides in nerve is correlated with electrophysiological nerve indices.…”