A feature of diabetic sensory polyneuropathy is that the earliest and most severe pathophysiology occurs in neurones with the longest axons. The aim of this study was to characterize a diabetes-induced neurodegenerative marker that was selective for sensory neurones with the longest axons. We studied alterations in calcium homeostasis since this occurs in other neurodegenerative diseases. Cultures of adult sensory neurones were prepared from control and streptozotocin (STZ)-diabetic rats, killed humanely according to UK legislation, treated with or without neurotrophin-3 (NT-3). Male Wistar rats (300 g) were made diabetic by a single intraperitoneal injection of STZ (55 mg kg^-1; Sigma). Age-matched and weight-matched control groups were also set up. Tail blood glucose was assayed 3 days after injection to confirm diabetes. All diabetic animals had blood glucose values higher than 30 mM. Sensory neurones from the lumber L4-L6 dorsal root ganglia (DRG) (these cells exhibit the longest axons in vivo) were compared with neurones from C1-L3 DRG. Fluorescent fura-2-based video-imaging was employed to measure calcium homeostasis. Significance was assessed by ANOVA.

STZ-diabetes, 8-14 weeks, induced a progressive increase in resting [Ca²⁺]_i, from 89 ± 9 nM (mean ± S.E.M.) in control animals to 237 ± 14 nM after 14 weeks of diabetes (P < 0.05) in L4-L6 DRG neurone cultures. Neurones cultured from C1-L3 ganglia and all non-neuronal cells were unaffected. Similar patterns were observed for stimulation-induced Ca²⁺ signalling. That is, the amplitude of depolarisation-induced Ca²⁺ transients decreased specifically in cells isolated from L4-L6 DRG, but not in cells from C1-L3 DRG. In addition, we found a profound retardation of the recovery of depolarisation-induced [Ca²⁺]_i transients in neurones isolated from diabetic rats. The slow down of [Ca²⁺]_i recovery was apparent in all cell groups, being the most prominent in neurones isolated from L4-L6 DRG after 8 weeks of diabetes. Experimental diabetes also caused a significant decrease in the number of L4-L6 neurones exhibiting [Ca²⁺]_i transients in response to stimulation of Ca²⁺ release from the ER stores (by 20 mM caffeine), which was paralleled by a decrease in the amplitude of caffeine-induced [Ca²⁺]_i responses. Conversely, the sensitivity of C1-L3 DRG neurones to caffeine was not affected by STZ diabetes. Systemic treatment of STZ-diabetic rats with NT-3 for 10 weeks reversed the described alterations of [Ca²⁺]_i homeostasis in L4-L6 DRG neurones.

These results show that altered calcium signalling may be an early predictor of diabetic sensory neuropathy and should encourage further work aimed at understanding the mechanistic basis of this observation.

This work was funded by grants from Diabetes UK (P.F. and A.V.) and the Juvenile Diabetes Research Foundation (P.F.). Tze-Jen Huang was supported by an Overseas Research Scholarship.