Endocrine Abstracts

JOINT858

Severe hypoglycemia (HPG) is a critical adverse effect of insulin therapy in diabetes, potentially leading to brain damage and cognitive impairment. This risk is one of the major hurdles for enough glucose control and can be a fear to patients and healthcare providers. Our study investigates the crosstalk between mitochondrial dynamics and neuroinflammation to understand hypoglycemic neuronal damage mechanisms and identify potential therapeutic interventions. Male C57BL/6 mice were fasted for 24 hours and HPG (below 20 mg/dl) was induced by intraperitoneal (i.p.) injection of insulin. HPG was induced for 5 hours and terminated by glucose solution i.p. injection. Mice were then sacrificed on day 1, 4, and 7. Unbiased screening in cortex and hippocampus area revealed the retrosplenial cortex (RSC) as vulnerable to HPG among other cortex regions, evidenced by elevated oxidative stress with 4-hydroxynonenal immunohistochemistry on day 7. Progressive increases in oxidative stress and apoptosis, analyzed by terminal deoxynucleotidyl transferase dUTP nick end labelling staining, were observed in the RSC. While mitochondrial fragmentation, examined through transmission electron microscopy, immunoblotting, and immunohistochemistry, and inflammatory activation with IL-1β expression level were already significantly increased at day 1 whereas TNF-α and IL-6 expression levels were unchanged. Treatment with mitochondrial fission inhibitor [mdivi-1] or IL-1 receptor antagonist (IL-1ra) effectively mitigated hypoglycemic neuronal damage. Notably, elevated mitochondrial fission was only significantly increased in neurons, not microglia and astrocytes, as analyzed by co-localization of phosphorylated dynamin-related protein, a marker of activated mitochondrial fission, with markers specific to neurons, microglia, and astrocytes. In vitro experiments with cell-type-specific regulation using transwell co-culture system revealed that preventing mitochondrial fragmentation with mdivi-1 in neuronal cells [SH-SY5Y], and inhibiting IL-1 signaling with IL-1ra in either microglial cells [BV-2] or SH-SY5Y significantly prevented hypoglycemic damage. Morris water maze assessments confirmed the protective effects of these interventions against spatial memory impairment induced by HPG. These results suggest that direct regulation of neuronal mitochondrial fission, combined with indirect modulation through the crosstalk between neuroinflammation with IL-1 signaling, could prevent hypoglycemic neuronal damage and spatial memory impairment.