Two Phases of Nitrergic Neuropathy in Streptozotocin-Induced Diabetic Rats

Abstract

The distinction between metabolic and structural changes occurring in autonomic neurons during diabetes has not been fully clarified. Here we demonstrate that nitric oxide synthase-containing (nitrergic) neurons innervating the penis and gastric pylorus of streptozotocin-induced diabetic rats undergo a selective degenerative process in two phases. In the first phase, nitrergic nerve fibers lose some of their neuronal nitric oxide synthase content and function. In the second phase, nitrergic degeneration takes place in the cell bodies in the ganglia, leading to complete loss of nitrergic function. The changes in the first phase are reversible with insulin replacement; however, the neurodegeneration in the second phase is irreversible. Neurodegeneration is due to apoptotic cell death in the ganglia, which is selective for the nitrergic neurones. Diabetes 52:2353-2362, 2003 D iabetes is a common disorder that leads to complications affecting the retina, kidney, vascular, gastrointestinal, peripheral, and autonomic nervous systems. The mechanism through which diabetic complications develop is unclear. The generally accepted view for most diabetic complications is that the disease is accompanied by metabolic changes in the affected organ that, in the long term, result in structural alterations. Thus, if insulin replacement is started before the structural lesions occur, then it should reverse the metabolic changes and prevent the development of the complication. Once the structural lesions occur, however, the process should become less reversible with insulin replacement. Although from early studies in vivo "a point of no return" had been suggested during the course of diabetic peripheral neuropathy, cardiomyopathy, and nephropathy (1-3), most of the subsequent work has concentrated either on cellular dysfunction or on cell death without addressing the distinction between metabolic and structural alterations.Nitric oxide (NO) is a well-characterized neurotransmitter in the central and peripheral nervous systems. In many tissues of the urogenital and gastrointestinal tract, NO mediates nonadrenergic noncholinergic (NANC) relaxant responses (4). Nerves that release NO are now known as nitrergic (5). NO is generated in these nerves by activation of the neuronal NO synthase (nNOS) and diffuses into the smooth muscle to activate the soluble guanylyl cyclase (sGC), producing an increase in the intracellular cyclic guanosine-3Ј, 5Ј-monophosphate concentration, leading to relaxation (6).Impaired nitrergic transmission has been shown to be responsible for erectile dysfunction and gastropathy in diabetes (7-12). To investigate the mechanisms by which the metabolic changes lead to structural disturbances in autonomic neuropathy, we studied the structure and function of nitrergic nerve fibers and neuronal cell bodies in the gastric pylorus, penis, and major pelvic ganglia in streptozotocin (STZ)-induced diabetic rats. In addition, we investigated the effect of different schedules of insulin treatment on this process. RESEARCH DE...

Synergistic action of advanced glycation end products and endogenous nitric oxide leads to neuronal apoptosis in vitro: A new insight into selective nitrergic neuropathy in diabetes

“…or vehicle (saline) as described previously [4,16]. Hyperglycaemia was defined as a non-fasting blood glucose concentration over 20 mmol/l.…”

“…SY5Y from European Cell Culture Collection; passages [13][14][15][16][17][18][19][20] were grown in DMEM/F12 mix (1:1) with glutamax-I (Life Technologies, Paisley, UK), 15% foetal calf serum, 10 mmol/l HEPES, 1% non-essential aminoacids and 1% antibiotics. Serum was withdrawn 24 h before any stimulation.…”

“…In the second phase the neurones undergo an apoptotic degenerative process and lose their nNOS content completely; this phase is irreversible by insulin treatment. Nitrergic responses are lost gradually throughout the two phases; this process is reversible by insulin treatment in the first but not in the second phase [16]. Interestingly in the second phase irreversible nitrergic degeneration as well as loss of nitrergic function occur even when the animals are made euglycaemic by treatment with insulin [16].…”

“…However, until recently it was not clear whether this decrease was due to degeneration of nitrergic nerves [4] or down-regulation of nNOS protein expression [5,15]. We have recently shown that [16] nitrergic neurones innervating the penis and gastric pylorus of diabetic rats go through a degenerative process in two phases: in the first phase the neuronal nitric oxide synthase (nNOS) content is decreased in the axons, but not in the cell bodies; this phase is reversible by insulin treatment and is not associated with neurodegenerative changes. In the second phase the neurones undergo an apoptotic degenerative process and lose their nNOS content completely; this phase is irreversible by insulin treatment.…”

“…Nitrergic responses are lost gradually throughout the two phases; this process is reversible by insulin treatment in the first but not in the second phase [16]. Interestingly in the second phase irreversible nitrergic degeneration as well as loss of nitrergic function occur even when the animals are made euglycaemic by treatment with insulin [16]. These findings prompted us to study factors other than high glucose that might lead to apoptotic neurodegeneration.…”