Reduced gap junctional communication among astrocytes in experimental diabetes: Contributions of altered connexin protein levels and oxidative–nitrosative modifications

Abstract

Experimental diabetes increases production of reactive oxygen-nitrogen species and inhibits astrocytic gap junctional communication in tissue culture and brain slices from streptozotocin (STZ)-diabetic rats by unidentified mechanisms. Relative connexin (Cx) protein levels were assessed by Western blotting using extracts from cultured astrocytes grown in high (25 mmol/L) or low (5.5 mmol/L) glucose for 2–3 weeks and STZ-diabetic rat brain. Chemiluminescent signals for diabetic samples were normalized to those of controls on the same blot and same protein load. Growth in high glucose did not alter relative Cx26 level, whereas Cx30 and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were reduced by ~30% and Cx43 increased ~1.9-fold. In the inferior colliculus of STZ-diabetic rats, Cx30 and Cx43 levels in three of four rats were half those of controls, whereas GAPDH and actin were unaffected. Diabetes did not affect levels of Cx30, Cx43, and GAPDH in cerebral cortex, but actin level rose 24%. Cx43 was predominantly phosphorylated in control and diabetic samples, so reduced dye transfer is not due to overall dephosphorylation of Cx43. Astrocytic growth in high glucose reduced dye-labeled area by 75%, but 10 min treatment with dithiothreitol restored normal dye transfer. In contrast, nitric oxide donors inhibited dye transfer among astrocytes grown in low glucose by 50–65% within 1h. Thus, modifications arising from oxidative-nitrosative stress, not altered connexin levels, may underlie reduced dye transfer among severely-hyperglycemic cultured astrocytes, whereas both oxidative-nitrosative stress and regionally-selective down-regulation of connexin protein content may affect gap junctional communication in brain of STZ-diabetic rats.

Astroglial Pentose Phosphate Pathway Rates in Response to High-Glucose Environments

“…If the adverse effects of high - glucose environments overcome the astroglial intrinsic protective mechanism in the long run, they may result in brain damage. Recent findings that experimental diabetes increases production of ROS–reactive nitrogen species and inhibits astrocytic gap junctional communication in tissue culture and brain slices from streptozotocin-diabetic rats suggest that astroglial dysfunction does, indeed, occur after longer period of disease duration irrespective of intrinsic self-defence mechanisms (Gandhi et al, 2010; Ball et al, 2011). Also, we observed increased immunostaining of N  ε -(carboxyethyl)lysine, one of the major AGEs, in astroglia that had been cultured in 23 mmol/l glucose for 6 weeks (Figure 11), supporting the above possibility.…”

High Glucose–Induced Downregulation of Connexin 30.2 Promotes Retinal Vascular Lesions: Implications for Diabetic Retinopathy

“…5 Other tissues such as corpus cavernosum exhibit reduced Cx43 in diabetes 20 and decreased expression of two connexins, Cx30 and Cx43, has been reported in the inferior colliculus of diabetic rats. 21 Most cells in general express more than one connexin. Since Cx30.2 has been reported to be expressed in the vasculature of different tissues, in this study, we determined whether Cx30.2 is expressed in the retinal vasculature and whether HG alters its expression and GJIC activity.…”

Connexin channel and its role in diabetic retinopathy

“…Impaired GJIC activity and Cx43 downregulation were observed in astrocytes and in brain slices of inferior colliculus in streptozotocin (STZ)-induced diabetic rat brains (Ball et al, 2011; Gandhi et al, 2010). These findings may have an effect on neurovascular unit function in diabetes.…”