Anne L. Carrington

Aldose reductase inhibitors and diabetic complications

Aldose reductase inhibitors impede flux of glucose through the sorbitol pathway in diabetes mellitus. They therefore reduce the accumulation of the pathway metabolites, sorbitol and fructose, reduce the impact of the flux on the cofactors used by the pathway and reduce other derived phenomena, such as osmotic stress and myo-inositol depletion. As drugs, their targets are the chronic complications of diabetes--neuropathy, retinopathy, nephropathy and vasculopathy. In experimental models there is proof of activity against biochemical, functional and structural defects in all of the involved tissues, but we await full clinical verification of this potential.

Nerve ischaemia in diabetic rats: time-course of development, effect of insulin treatment plus comparison of streptozotocin and BB models.

SummaryThis study sought to determine the timecourse of development of reduced nerve laser Doppler flux in experimental diabetes and the effect on this anomaly of insulin treatment. In addition, we aimed to compare nerve laser Doppler flux in streptozotocin-and genetically-diabetic BB rat models. Sciatic nerve laser Doppler flux in diabetic rats was variable during the 2 days following streptozotocin injection; from day 4, when the measurement was 80% of control, fluxes fell steadily and formed a plateau at 40% of control values after 4 weeks of diabetes. In a second study, using rats with 4-week streptozotocin-diabetes, sciatic nerve laser Doppler flux was reduced to 44% of the value measured in control rats. Treatment of a parallel group of diabetic rats with insulin, by sustained release implants, prevented this ischaemia, so that nerve laser Doppler flux was 91% of controls. Nerve Doppler flux in BB rats with 6-week genetic diabetes was 57% of a control (non-diabetic) BB group. There were no differences in mean arterial pressures between control and diabetic rats in any of the studies. Heart rates of control and insulin-treated diabetic animals were higher than those of the untreated diabetic group; in the other studies heart rates of diabetic animals were numerically lower than controls, but not significantly so. These observations suggest that sciatic nerves of rats with short-term diabetes, whether induced with streptozotocin or of genetic origin, are markedly ischaemic and that this ischaemia in streptozotocindiabetes is evident within a week of diabetes onset, forms a plateau after 4 weeks and is maintained for at least 2 months. The findings also indicate that treatment of short-term diabetes with insulin can prevent nerve ischaemia.

Effects of treatment with myo-inositol or its 1,2,6-trisphosphate (PP56) on nerve conduction in streptozotocin-diabetes

Nerve conduction abnormalities in peripheral nerves from diabetic patients may be early indicators for the future development of symptomatic neuropathy. In this study, three weeks of experimental diabetes in the rat caused a significant decrease in motor nerve conduction velocity measured in vivo (45.3 +/- 3.6 m/s; mean +/- S.D.) compared to controls (57.7 +/- 4.5 m/s). myo-Inositol administration to diabetic rats (500 mg/rat per day) for the duration of the study, partially prevented this decrease (50 +/- 4.4 m/s). An analogue of myo-inositol, PP56 (D-myo-inositol-1,2,6-trisphosphate), at a dose of 24 mg/rat per day completely prevented this reduction in diabetic rats (53.4 +/- 5.8 m/s). Resistance to hypoxic conduction block was determined in vitro in endoneurial preparations and was assessed as the decline in compound action potential amplitude over a 40 min period of hypoxia. Compound action potential amplitude (as % of initial value +/- S.D.) was significantly greater in diabetic preparations compared with controls at 40 min of hypoxia (76.1 +/- 9.1 vs. 54.8 +/- 14.7 respectively). Treatment to diabetic rats with myo-inositol did not significantly affect this value (79.9 +/- 16.6) but PP56 treatment partially prevented the increased resistance to hypoxic conduction block (69.4 +/- 16.0). This study demonstrates that these acute abnormalities of nerve function in early experimental diabetes may be attenuated by the administration of PP56, possibly acting via a vascular mechanism.

Enteric neuropeptides in streptozotocin-diabetic rats ; effects of insulin and aldose reductase inhibition

The aim of the present study was to determine whether diabetes-induced changes in the distribution of enteric neuropeptides, could be prevented in 12-week streptozotocin-diabetic rats, by rigorous control of glycaemia, using daily adminstration of insulin, or an aldose reductase inhibitor (ponalrestat). The pattern of distribution of nerve fibres and cell bodies, containing immunoreactive vasoactive intestinal polypeptide (VIP), galanin (GAL), calcitonin gene-related peptide (CGRP) and substance P was examined in the myenteric plexus of ileum from control, untreated diabetic, insulin-treated diabetic and aldose reductase inhibitor-treated diabetic rats. The increase in VIP- and GAL-like immunoreactivity, seen in the myenteric plexus of untreated diabetic rat ileum, was not present in the myenteric plexus of ileum from insulin- and aldose reductase inhibitor-treated diabetic rats. With CGRP-like immunoreactive fibres, there was a clear decrease in the ileum of untreated diabetic rats. This was prevented by insulin treatment, but aldose reductase inhibitor treatment had no effect. No alterations in substance P-like immunoreactivity were seen in the myenteric plexus of ileum from any of the groups investigated. Generally, the similarity of effect of ponalrestat and insulin on VIP and galanin expression in this study supports a primary effect of insulin via glycaemic control. The dissimilarity of the effect of the two treatments on CGRP expression may imply a neurotrophic effect of insulin, although there are certainly consequences of hyperglycaemia other than exaggerated flux through the polyol pathway.

Essential fatty acid treatment prevents nerve ischaemia and associated conduction anomalies in rats with experimental diabetes mellitus

SummaryIn rats with 6 weeks streptozotocin-diabetes there was a 53% reduction in sciatic nerve laser Doppler flux compared to controls (p<0.01). Treatment of a parallel group of diabetic rats with evening primrose oil, by dietary admixture throughout the protocol, prevented this ischaemia (Doppler flux was 91% of evening primrose oil-treated controls and was not significantly different). There were no differences in systemic arterial pressure. In another experiment evening primrose oil markedly antagonised the development of exaggerated resistance to anoxic conduction failure in sciatic nerves from diabetic rats. The resistance to anoxia of nerves from non-diabetic rats was also reduced by evening primrose oil. These observations suggest that the sciatic nerves of diabetic rats with short-term streptozotocin-diabetes are markedly ischaemic and that this ischaemia is involved in the development of increased resistance to anoxic/ischaemic conduction failure in diabetic nerve. The findings also promote evening primrose oil as a potential treatment to prevent nerve ischaemia.

Prostacyclin release in experimental diabetes: Effects of evening primrose oil

Alterations in release of endothelium-derived vasomotor agents could underlie microvascular and neuropathic complications in diabetes. This study examined release of the potent vasodilator prostacyclin, measured as immunoreactive 6-keto prostaglandin F1 alpha, from rat lung, kidney and peripheral nerve. Tissues were taken from control and streptozotocin-diabetic rats which had been treated for 8 weeks with either evening primrose oil (EPO) or, as a control for lipid intake, coconut oil (CO). Lung and kidney slices were incubated in the presence of acetylcholine (ACh), the calcium ionophore 4-Br-A23187, arachidonic acid (AA) or without agonist (basal). Segments of sciatic nerve, with their epineuria punctured, were incubated with or without 4-Br-A23187. Basal prostacyclin release from the lung was significantly higher in rats treated with EPO irrespective of diabetic state (increased by 60% in controls and by 77% in diabetics). Levels were reduced in CO-diabetics compared to EPO-controls (53% reduction) and CO-controls (30% reduction), although this did not reach statistical significance in the latter. Basal prostacyclin release was also significantly reduced in the kidney from CO-diabetics (40% reduction compared to CO-controls and 56% reduction compared to EPO-controls). In the presence of AA, lung prostacyclin release was significantly lower in CO-diabetic rats compared to all other groups (40% reduction compared to EPO-diabetics and 60% compared to both control groups) but there were no differences in renal release between any group. Prostacyclin release by nerves from CO-diabetic rats was significantly reduced (by 91-93%) compared to all other groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Aldose reductase inhibition with imirestat — effects on impulse conduction and insulin-stimulation of Na+/K+-adenosine triphosphatase activity in sciatic nerves of streptozotocin-diabetic rats

SummaryThis study describes reduced motor nerve conduction velocity and increased resistance to hypoxia-induced conduction failure in sciatic nerves of rats after four weeks of streptozotocin-induced diabetes (both effects were significant at p <0.05). These changes occurred in the absence of any deficit in the steady-state ouabain-sensitive adenosine triphosphatase (ATPase) activity of sciatic nerve endoneurial homogenates. The addition of 10 nmol/l insulin to endoneurial homogenates from control animals resulted in a 34% increase in ouabain-sensitive ATPase activity and a 19% reduction in ouabain-insensitive ATPase activity (both p <0.01). This stimulation of ouabain-sensitive ATPase activity by insulin did not occur in homogenates from diabetic rats. Treating diabetic rats daily with the aldose reductase inhibitor, imirestat (1 mg/kg) improved nerve conduction velocity (p <0.05) but was without effect upon the resistance to hypoxic conduction blockade or the deficit in insulin-stimulated oubain-sensitive ATPase activity. These data suggest that in streptozotocin-diabetic rats the functional disorders of reduced motor nerve conduction velocity and increased resistance to hypoxic conduction blockade do not share a common aetiology and that impaired nerve conduction is not related to reduced maximal potential oubain-sensitive ATPase activity.

Reduced nerve blood flow in diabetic rats: relationship to nitric oxide production and inhibition of aldose reductase

This study examined links between impaired nitric oxide production in the sciatic endoneurium, nerve blood flow, and polyol pathway flux, to test the hypothesis that reduced nerve blood flow might be compromised by competition for NADPH between aldose reductase and nitric oxide synthase. Sciatic nerves of streptozotocin‐diabetic rats showed reduced laser Doppler flux (by 51 % or 63 %; both p<0.05)—indicative of reduced nerve blood flow—and reduced motor nerve conduction velocity (17 % in two experiments; p<0.05). Acute interruption of nitric oxide production in the sciatic nerves of control rats, via endoneurial injection of Nω‐nitro‐D‐arginine methyl ester ( L‐NAME), caused a local reduction (of 64 %; p<0.001) in nerve Doppler flux. This was reversed by either L‐arginine or sodium nitroprusside. The response to L‐NAME was greatly reduced in diabetic rats (only 22 % reduction; p<0.01), though both L‐arginine and SNP caused marked increases in flux. Chronic inhibition of aldose reductase in diabetic rats (with either sorbinil or imirestat at a range of doses) had little effect on resting sciatic nerve Doppler flux, though both inhibitors normalized conduction velocity. Both aldose reductase inhibitors reduced sorbitol pathway intermediates in a dose‐related manner. These findings do not support the proposition that aldose reductase inhibitors normalise conduction velocity by mechanisms dependent upon either normalization of endoneurial nitric oxide or nerve blood flow. Instead, a mechanism based upon more direct effects on axon or Schwann cell function is favoured.

Resistance to hypoxic conduction block in sciatic nerves of rats with streptozotocin-induced diabetes mellitus

This study describes the electrophysiological responses of endoneurial preparations derived from rat sciatic nerve to acute hypoxia in vitro. Preparations from control rats exhibited a marked decline in compound action potential (CAP) amplitude coupled with an increase in latency, during 40 minutes exposure to 8% O2. In contrast, preparations from 4 week streptozotocin-diabetic rats showed a greatly reduced decline in CAP amplitude, with an increase in latency. Twice-daily insulin treatment of diabetic rats resulted in a pattern of CAP amplitude decline that initially resembled that of untreated diabetics but by 40 min was similar to controls, with latency again increasing during hypoxia. Nerves were also maintained in 25 mmol/l glucose, rather than the 5 mmol/l glucose of the above studies. Under such conditions the performance of nerves from diabetic rats was unaltered. Nerves from control rats exhibited an initial resistance to hypoxia but by 40 min CAP had declined to values of control rats maintained in 5 mmol/l glucose. An increase in latency during hypoxia was also noted in preparations from control or diabetic rats maintained in 25 mmol/l glucose. The maintenance of CAP amplitude during hypoxia by diabetic preparations is initially related to increased substrate availability, with an additional component that is not related to external glucose levels in vitro, and is absent after insulin treatment of diabetic rats.

Increased resitance to hypoxic conduction block in sciatic nerves of diabetic rats: Effects of extracellular glucose concentration and of aldose reductase inhibition

This study examined the effect of the aldose reductase inhibitor (ARI), ponalrestat, on decreased motor nerve conduction velocity (MNCV) and increased resistance to hypoxic conduction block (RHCB) in diabetic rats. The effects of 5 mmol/L, and 25 mmol/L glucose on RHCB were also determined. Twenty streptozotocin-diabetic rats formed two groups; untreated and ponalrestat-treated (300 mg/kg diet/day); 10 non-diabetic rats acted as controls. MNCV was measured in vivo after 4 weeks of diabetes +/- treatment in the sciatic/tibialis system and rats were killed 48-72 h later. The median nerves were removed and assayed for polyol pathway metabolites by gas chromatography. The sciatic nerves were dissected to form endoneurial preparations for the recording of compound action potentials (CAPs) in vitro and maintained in media with either 5 (standard) or 25 (high) mmol/L glucose and initially gassed with 95% O2/5% CO2. Oxygen content was then reduced to 8% for 40 min to study the effect of this period of hypoxia on CAP amplitude. MNCV (m/s +/- SD) in diabetic rats (43.86 +/- 4.86) was decreased compared to controls (52.24 +/- 6.59) and this decrease was absent in the ARI-treated group (52.24 +/- 6.90). The decline in CAP amplitude during a 40-min hypoxic period was greater in controls than in diabetics. Ponalrestat treatment caused a decline which was mid-way between these two in standard medium and closer to that seen in control preparations in high glucose medium. These findings give further support to the involvement of the sorbitol pathway in the development of the acute MNCV deficit in diabetic rats and indicate that it may have a partial role in the development of increased resistance to hypoxic conduction block in peripheral nerves.