No abstract

The contrast between resistance to ischemia and ischemic lesions in peripheral nerves of diabetic patients was explored by in vitro experiments. Isolated and desheathed rat peroneal nerves were incubated in the following solutions with different glucose availability: 1) 25 mM glucose, 2) 2.5 mM glucose, and 3) 2.5 mM glucose plus 10 mM 2-deoxy-D-glucose. Additionally, the buffering power of all of these solutions was modified. Compound nerve action potential (CNAP), extracellular pH, and extracellular potassium activity (aKe) were measured simultaneously before, during, and after a period of 30 min of anoxia. An increase in glucose availability led to a slower decline in CNAP and to a smaller rise in aKe during anoxia. This resistance to anoxia was accompanied by an enhanced extracellular acidosis. Postanoxic recovery of CNAP was always complete in 25 mM HCO3(-)-buffered solutions. In 5 mM HCO3- and in HCO3(-)-free solutions, however, nerves incubated in 25 mM glucose did not recover functionally after anoxia, whereas nerves bathed in solutions 2 or 3 showed a complete restitution of CNAP. We conclude that high glucose availability and low PO2 in the combination with decreased buffering power and/or inhibition of HCO3(-)-dependent pH regulation mechanisms may damage peripheral mammalian nerves due to a pronounced intracellular acidosis.

1. Electrotonic responses to 150 ms current pulses were recorded from isolated rat dorsal roots incubated for at least 3 h with either normal (5 mM) or high (25 mM) D-glucose solutions, and with either normal (25 mM) or low (5 mM) bicarbonate concentrations. 2. On replacement of 02 by N2 for 50 min, all the roots depolarized, but the changes in electrotonus differed systematically. With normal glucose, the depolarization was accompanied by an increase in input conductance. In contrast, for the hyperglyeaemic roots the depolarization was slower and accompanied by a fall in input conductance which was exacerbated in low bicarbonate concentrations. 3. The changes induced by hyperglycaemic hypoxia in low bicarbonate could be mimicked by exposure of the roots either to 100% CO2 or to a combination of 3 mm tetraethylammonium chloride and 3 mm 4-aminopyridine, to block both fast and slow potassium channels. 4. These results indicate that the primary mechanism of hypoxic depolarization of these sensory axons is altered by hyperglycaemia. In normoglycaemia, the changes in electrotonus are consistent with an increase in axonal potassium conductance. The block of potassium channels seen in hyperglycaemic hypoxia is attributed to intra-axonal acidification by anaerobic glycolysis and may contribute to the pathogenesis of diabetic neuropathy.

We explore whether the prevalence of sensory deficits in diabetic neuropathy can be explained by diffuse endoneurial hypoxia. Isolated ventral and dorsal rat spinal roots incubated in 2.5 or 25 mM extracellular glucose were transiently exposed to hypoxia (30 min) in a solution of low buffering power. Compound nerve action potentials and extracellular direct current potentials were continuously recorded before, during, and after hypoxia. In both ventral and dorsal roots incubated in 2.5 mM glucose, sensitivity to hypoxia and posthypoxic recovery were similar. In contrast, hypoxia in 25 mM glucose preferentially induced electrophysiological damage in dorsal roots as indicated by a lack of posthypoxic recovery. This observation was not made in the presence of 25 mM bicarbonate, which suggests involvement of nerve acidosis. In conclusion, the different sensitivity of sensory and motor fibers to hyperglycemic hypoxia supports the hypothesis that hypoxia has an important role in the pathogenesis of diabetic neuropathy.

SUMMARY1. The effects of hyperglyeaemic hypoxia (a condition possibly involved in the pathogenesis of diabetic neuropathy) on the depolarizing after-potential and the potassium conductance of myelinated rat spinal root axons were investigated using electrophysiological recordings from intact spinal roots and from excised, inside-out axonal membrane patches.2. Isolated spinal roots were exposed to hypoxia in solutions containing normal or high glucose concentrations. The depolarizing after-potential of compound action potentials was only enhanced in spinal roots exposed to hyperglycaemic (25 mM Dglucose) hypoxia. A maximal effect was seen in bathing solutions with low buffering power.3. The depolarizing after-potential was also enhanced by cytoplasmic acidification after replacement of 10-30 mm chloride in the bathing solution by propionate.4. Multi-channel current recordings from excised, inside-out axonal membrane patches were used to study the effects of cytoplasmic acidification on voltagedependent K+ conductances with fast (F channels) and intermediate (I channels) kinetics of deactivation.5. F channels were blocked by small changes in cytoplasmic pH (50 % inhibition at pH 6 9). I channels were much less sensitive to intra-axonal acidification.6. In conclusion, our data show that hyperglycaemic hypoxia enhances the depolarizing after-potential in peripheral rat axons. The underlying mechanism seems to be an inhibition of a fast, voltage-dependent axonal K+ conductance by cytoplasmic acidification. This alteration in membrane conductance may contribute to positive symptoms in diabetic neuropathy.