Insulin-induced recurrent hypoglycemia exacerbates diabetic brain mitochondrial dysfunction and oxidative imbalance

Susana Cardoso, Renato X. Santos, Sónia C. Correia, Cristina Carvalho, Maria S. Santos, Inês Baldeiras, Catarina R. Oliveira, Paula I. Moreira (Corresponding Author)

Research output: Contribution to journalArticle

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Abstract

Intensive insulin therapy can prevent or slow the progression of long-term diabetes complications but, at the same time, it increases the risk for episodes of severe hypoglycemia. In our study, we used a protocol intended to mimic the levels of blood glucose that occur in type 1 diabetic patients under an intensive insulin therapy. Streptozotocin (STZ)-induced diabetic rats were treated subcutaneously with twice-daily insulin injections for 2 weeks to induce hypoglycemic episodes. Brain cortical and hippocampal mitochondria were isolated and mitochondrial bioenergetics (respiratory chain and phosphorylation system) and oxidative status parameters (malondialdehyde (MDA) levels, mitochondrial aconitase activity and enzymatic and non-enzymatic antioxidant defenses) were analyzed. The protein levels of synaptophysin, a marker of synaptic integrity, and caspase 9 activity were also evaluated in cortical and hippocampal homogenates. Brain cortical mitochondria isolated from hyper- and recurrent hypoglycemic animals presented higher levels of MDA and α-tocopherol together with an increased glutathione disulfide reductase activity, lower manganese superoxide dismutase (MnSOD) activity and glutathione-to-glutathione disulfide (GSH/GSSG) ratio. No significant alterations were found in cortical mitochondrial respiratory chain and oxidative phosphorylation system. Hippocampal mitochondria from both experimental groups presented an impaired oxidative phosphorylation system characterized by a decreased mitochondrial energization potential and ATP levels and higher repolarization lag phase. In addition, higher MDA levels and decreased GSH/GSSG, α-tocopherol levels, and aconitase, glutathione peroxidase and MnSOD activities were observed in both groups of animals. Hippocampal mitochondria from recurrent hypoglycemic animals also showed an impairment of the respiratory chain characterized by a lower state 3 of respiration, respiratory control ratio and ADP/O index, and a higher state 4 of respiration. Additionally, a non-statistically significant decrease in synaptophysin protein levels was observed in cortical homogenates from recurrent hypoglycemic rats as well as in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic rats. An increase in caspase 9 activity was also observed in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic animals. Our results show that mitochondrial dysfunction induced by long-term hyperglycemic effects is exacerbated by recurrent hypoglycemia, which may compromise the function and integrity of brain cells.
Original languageEnglish
Pages (from-to)1-12
Number of pages12
JournalNeurobiology of Disease
Volume49
Early online date24 Aug 2012
DOIs
Publication statusPublished - Jan 2013

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Hypoglycemia
Hypoglycemic Agents
Insulin
Glutathione Disulfide
Oxidative Phosphorylation
Mitochondria
Brain
Electron Transport
Malondialdehyde
Aconitate Hydratase
Synaptophysin
Tocopherols
Caspase 9
Superoxide Dismutase
Respiration
Glutathione Reductase
Diabetes Complications
Streptozocin
Glutathione Peroxidase
Adenosine Diphosphate

Keywords

  • Cortex
  • Hippocampus
  • Long-term hyperglycemia
  • Mitochondria
  • Neurodegeneration
  • Oxidative stress
  • Recurrent hypoglycemia

Cite this

Insulin-induced recurrent hypoglycemia exacerbates diabetic brain mitochondrial dysfunction and oxidative imbalance. / Cardoso, Susana; Santos, Renato X.; Correia, Sónia C.; Carvalho, Cristina; Santos, Maria S.; Baldeiras, Inês; Oliveira, Catarina R.; Moreira, Paula I. (Corresponding Author).

In: Neurobiology of Disease, Vol. 49, 01.2013, p. 1-12.

Research output: Contribution to journalArticle

Cardoso, Susana ; Santos, Renato X. ; Correia, Sónia C. ; Carvalho, Cristina ; Santos, Maria S. ; Baldeiras, Inês ; Oliveira, Catarina R. ; Moreira, Paula I. / Insulin-induced recurrent hypoglycemia exacerbates diabetic brain mitochondrial dysfunction and oxidative imbalance. In: Neurobiology of Disease. 2013 ; Vol. 49. pp. 1-12.
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abstract = "Intensive insulin therapy can prevent or slow the progression of long-term diabetes complications but, at the same time, it increases the risk for episodes of severe hypoglycemia. In our study, we used a protocol intended to mimic the levels of blood glucose that occur in type 1 diabetic patients under an intensive insulin therapy. Streptozotocin (STZ)-induced diabetic rats were treated subcutaneously with twice-daily insulin injections for 2 weeks to induce hypoglycemic episodes. Brain cortical and hippocampal mitochondria were isolated and mitochondrial bioenergetics (respiratory chain and phosphorylation system) and oxidative status parameters (malondialdehyde (MDA) levels, mitochondrial aconitase activity and enzymatic and non-enzymatic antioxidant defenses) were analyzed. The protein levels of synaptophysin, a marker of synaptic integrity, and caspase 9 activity were also evaluated in cortical and hippocampal homogenates. Brain cortical mitochondria isolated from hyper- and recurrent hypoglycemic animals presented higher levels of MDA and α-tocopherol together with an increased glutathione disulfide reductase activity, lower manganese superoxide dismutase (MnSOD) activity and glutathione-to-glutathione disulfide (GSH/GSSG) ratio. No significant alterations were found in cortical mitochondrial respiratory chain and oxidative phosphorylation system. Hippocampal mitochondria from both experimental groups presented an impaired oxidative phosphorylation system characterized by a decreased mitochondrial energization potential and ATP levels and higher repolarization lag phase. In addition, higher MDA levels and decreased GSH/GSSG, α-tocopherol levels, and aconitase, glutathione peroxidase and MnSOD activities were observed in both groups of animals. Hippocampal mitochondria from recurrent hypoglycemic animals also showed an impairment of the respiratory chain characterized by a lower state 3 of respiration, respiratory control ratio and ADP/O index, and a higher state 4 of respiration. Additionally, a non-statistically significant decrease in synaptophysin protein levels was observed in cortical homogenates from recurrent hypoglycemic rats as well as in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic rats. An increase in caspase 9 activity was also observed in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic animals. Our results show that mitochondrial dysfunction induced by long-term hyperglycemic effects is exacerbated by recurrent hypoglycemia, which may compromise the function and integrity of brain cells.",
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AU - Santos, Maria S.

AU - Baldeiras, Inês

AU - Oliveira, Catarina R.

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N1 - Acknowledgments Susana Cardoso has a PhD fellowship from the Portuguese Foundation for Science and Technology (SFRH/BD/43968/2008). We thank Dr. Paula Agostinho for kindly providing the synaptophysin antibody.

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N2 - Intensive insulin therapy can prevent or slow the progression of long-term diabetes complications but, at the same time, it increases the risk for episodes of severe hypoglycemia. In our study, we used a protocol intended to mimic the levels of blood glucose that occur in type 1 diabetic patients under an intensive insulin therapy. Streptozotocin (STZ)-induced diabetic rats were treated subcutaneously with twice-daily insulin injections for 2 weeks to induce hypoglycemic episodes. Brain cortical and hippocampal mitochondria were isolated and mitochondrial bioenergetics (respiratory chain and phosphorylation system) and oxidative status parameters (malondialdehyde (MDA) levels, mitochondrial aconitase activity and enzymatic and non-enzymatic antioxidant defenses) were analyzed. The protein levels of synaptophysin, a marker of synaptic integrity, and caspase 9 activity were also evaluated in cortical and hippocampal homogenates. Brain cortical mitochondria isolated from hyper- and recurrent hypoglycemic animals presented higher levels of MDA and α-tocopherol together with an increased glutathione disulfide reductase activity, lower manganese superoxide dismutase (MnSOD) activity and glutathione-to-glutathione disulfide (GSH/GSSG) ratio. No significant alterations were found in cortical mitochondrial respiratory chain and oxidative phosphorylation system. Hippocampal mitochondria from both experimental groups presented an impaired oxidative phosphorylation system characterized by a decreased mitochondrial energization potential and ATP levels and higher repolarization lag phase. In addition, higher MDA levels and decreased GSH/GSSG, α-tocopherol levels, and aconitase, glutathione peroxidase and MnSOD activities were observed in both groups of animals. Hippocampal mitochondria from recurrent hypoglycemic animals also showed an impairment of the respiratory chain characterized by a lower state 3 of respiration, respiratory control ratio and ADP/O index, and a higher state 4 of respiration. Additionally, a non-statistically significant decrease in synaptophysin protein levels was observed in cortical homogenates from recurrent hypoglycemic rats as well as in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic rats. An increase in caspase 9 activity was also observed in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic animals. Our results show that mitochondrial dysfunction induced by long-term hyperglycemic effects is exacerbated by recurrent hypoglycemia, which may compromise the function and integrity of brain cells.

AB - Intensive insulin therapy can prevent or slow the progression of long-term diabetes complications but, at the same time, it increases the risk for episodes of severe hypoglycemia. In our study, we used a protocol intended to mimic the levels of blood glucose that occur in type 1 diabetic patients under an intensive insulin therapy. Streptozotocin (STZ)-induced diabetic rats were treated subcutaneously with twice-daily insulin injections for 2 weeks to induce hypoglycemic episodes. Brain cortical and hippocampal mitochondria were isolated and mitochondrial bioenergetics (respiratory chain and phosphorylation system) and oxidative status parameters (malondialdehyde (MDA) levels, mitochondrial aconitase activity and enzymatic and non-enzymatic antioxidant defenses) were analyzed. The protein levels of synaptophysin, a marker of synaptic integrity, and caspase 9 activity were also evaluated in cortical and hippocampal homogenates. Brain cortical mitochondria isolated from hyper- and recurrent hypoglycemic animals presented higher levels of MDA and α-tocopherol together with an increased glutathione disulfide reductase activity, lower manganese superoxide dismutase (MnSOD) activity and glutathione-to-glutathione disulfide (GSH/GSSG) ratio. No significant alterations were found in cortical mitochondrial respiratory chain and oxidative phosphorylation system. Hippocampal mitochondria from both experimental groups presented an impaired oxidative phosphorylation system characterized by a decreased mitochondrial energization potential and ATP levels and higher repolarization lag phase. In addition, higher MDA levels and decreased GSH/GSSG, α-tocopherol levels, and aconitase, glutathione peroxidase and MnSOD activities were observed in both groups of animals. Hippocampal mitochondria from recurrent hypoglycemic animals also showed an impairment of the respiratory chain characterized by a lower state 3 of respiration, respiratory control ratio and ADP/O index, and a higher state 4 of respiration. Additionally, a non-statistically significant decrease in synaptophysin protein levels was observed in cortical homogenates from recurrent hypoglycemic rats as well as in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic rats. An increase in caspase 9 activity was also observed in hippocampal homogenates from hyperglycemic and recurrent hypoglycemic animals. Our results show that mitochondrial dysfunction induced by long-term hyperglycemic effects is exacerbated by recurrent hypoglycemia, which may compromise the function and integrity of brain cells.

KW - Cortex

KW - Hippocampus

KW - Long-term hyperglycemia

KW - Mitochondria

KW - Neurodegeneration

KW - Oxidative stress

KW - Recurrent hypoglycemia

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