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Copper loading may affect rat neurobehaviour by impairing mitochondria-associated endoplasmic reticulum membranes in
Zhengzhe Sun1, Shan Jin2, Xiang Fang3,4
1The Second Affiliated Hospital of Anhui University of Traditional Chinese Medicine, Hefei, Anhui, China.
Background:
To observe the effects of copper sulfate (CuSO4)-induced copper loading on neurobehaviour, mitochondria-associated endoplasmic reticulum membranes (MAMs) and related regulatory proteins in the hippocampal CA1 region of Sprague-Dawley (SD) rats.
Methods:
Forty SD male rats were randomly divided into control and copper loading groups of 20 rats each. The control group rats were fed with normal feed and water; rats in the copper loading group were fed high copper feed (containing 1g/kg of CuSO4) and CuSO4 deionized water (concentration of 0.185%). After 12 weeks of rearing, the morris water maze (MWM) task and novel object recognition (NOR) test were conducted to compare the neurobehavioral characteristics of the two groups of rats. Morphological changes of neuronal MAMs in the hippocampal CA1 region of copper-loaded rats were observed using a transmission electron microscope (TEM) and immunofluorescence double-labelling techniques. Western-blot analysis was used to detect the expression of MAMs proteins VDAC1, IP3R, GRP75 and Mfn2.
Results:
The results revealed that rats in the copper-loading group had significantly prolonged escape latency and reduced number of platform crossings in the MWM task (p < 0.01). The percentage of novel objects explored (also known as the Discrimination Ratio, DR) and the discrimination index (DI) were significantly reduced in the NOR test (p < 0.01). In addition, electron microscopy shows increased disruption of neuronal endoplasmic reticulum (ER)-mitochondrion coupling in the hippocampal CA1 region of rats in the copper-loading group (p < 0.05), and the percentage of MAMs in mitochondrial circumference decreased (p < 0.05), the colocalization coefficients between the ER and mitochondria was significantly reduced (p < 0.05). Moreover, the protein expression levels of VDAC1, IP3R, and GRP75 in rat hippocampal tissue were detected to be significantly increased (p < 0.01), while the protein expression level of Mfn2 was significantly decreased (p < 0.01).
Conclusions:
In this study, it is speculated that the neurobehavioral changes in rats may be related to the increased expression levels of the MAMs proteins VDAC1, IP3R, and GRP75, the reduced expression level of Mfn2, and the disruption of the structural integrity of MAMs in the hippocampal CA1 region of rats caused by copper loading.
Insights
Copper sulfate exposure in rats impaired neurobehavior and disrupted mitochondria-associated endoplasmic reticulum membranes (MAMs) in the hippocampus. These changes were linked to altered MAMs protein levels, suggesting a mechanism for copper-induced neurotoxicity.
Area of Science:
- Neuroscience
- Cell Biology
- Toxicology
Background:
- Copper is essential but toxic in excess.
- Copper sulfate (CuSO4) exposure can induce copper loading.
- The hippocampus is vulnerable to neurotoxic insults.
Purpose of the Study:
- To investigate the impact of copper loading on neurobehavior in rats.
- To examine alterations in mitochondria-associated endoplasmic reticulum membranes (MAMs) and associated proteins in the hippocampus.
- To correlate neurobehavioral changes with MAMs integrity and protein expression.
Main Methods:
- Adult male Sprague-Dawley rats were divided into control and copper-loaded groups.
- Copper loading was induced via CuSO4 in feed and water for 12 weeks.
- Neurobehavior was assessed using the Morris Water Maze (MWM) and Novel Object Recognition (NOR) tests.
- Hippocampal CA1 region MAMs morphology was analyzed via transmission electron microscopy (TEM).
- Expression of MAMs proteins (VDAC1, IP3R, GRP75, Mfn2) was quantified using Western blot analysis.
Main Results:
- Copper-loaded rats exhibited significantly impaired performance in MWM and NOR tests.
- TEM revealed disrupted ER-mitochondrion coupling and reduced MAMs in the hippocampal CA1 region.
- Protein analysis showed increased levels of VDAC1, IP3R, and GRP75, and decreased Mfn2.
- These structural and molecular changes were statistically significant (p < 0.01).
Conclusions:
- Copper loading induces significant neurobehavioral deficits in rats.
- Disruption of MAMs integrity and altered expression of key MAMs proteins are associated with copper-induced neurotoxicity.
- These findings suggest that MAMs dysfunction plays a critical role in the neurotoxic effects of excess copper.
