Research reportChronic social isolation decreases glutamate and glutamine levels and induces oxidative stress in the rat hippocampus
Introduction
Social isolation (SI) rearing of rodents is a developmental manipulation in which an animal is singly housed upon weaning. This manipulation is commonly compared with the psychological stressors in humans as it produces several behavioral outcomes similar to those observed in humans with early life stress. The SI-induced behavioral outcomes include anxiety/depression-like behaviors [2], [4], [6], [9], [51], [54] and deficits in social interaction [33], [34], [52], [55], prepulse inhibition (PPI) of the acoustic startle reflex [12], [33], sensory gating and mismatch negativity (MMN)-like responses [50], and in cognition and memory [13], [15], [36]. Of the SI-induced behavioral outcomes, deficits in PPI, MMN deficits [43], and increased preservative behavior resemble some of the symptomatology of schizophrenia. Moreover, the behavioral effects of isolation rearing on rats were ameliorated by the antipsychotic clozapine [33], [34].
In quest of the neural substrate of the SI-induced behavioral deficits, previous animal studies compared some specific brain regions of isolation-reared animals with normal controls in terms of the indices relevant to the dopaminergic and glutamatergic systems, both of which are implicated in schizophrenia [7], [42]. For example, isolation-reared rats showed higher levels of basal extracellular dopamine in the stratum [16]. Other studies tried to find glutamatergic alterations in isolation-reared animals. For example, an earlier in situ hybridization study noted that mRNA of the N-methyl-d-aspartate (NMDA) receptor subunit NR1A was decreased in the hippocampus of isolation-reared rats compared with their socially housed counterparts [17]. Also, SI was reported to up-regulate NR2A expression in the prefrontal cortex (PFC) of rats [47], increase NMDA receptor binding in frontal cortex [46], and stimulate the mRNA expression of NR2A and NR2B in the hippocampus [55]. In a more recent study, ‘SI’ increased the binding activity of the group II metabotropic glutamate receptor (mGluR2/3) in the PFC and hippocampus of mice [24]. These previous data suggest the existence of glutamatergic alterations in brains of isolation reared rodents. However, there is little information available regarding the brain metabolic profile measured in living animals using non-invasive neuroimaging methods.
The SI-induced behavioral outcomes were also associated with cellular oxidative stress. In mice, SI for 6–8 weeks induced oxidative damage in the brain by enhancing the production of nitric oxide and depleting brain glutathione content, an endogenous antioxidant [21]. In rats, chronic SI compromised the activities of both glutathione peroxidase (GPx) and catalase in the hippocampus [11]. According to the authors, the mechanism for the SI-induced oxidative imbalance in the hippocampus is likely due to poor systemic energy conditions set by this stressor.
The aims of this study were to detect possible changes in levels of glutamate and glutamine caused by SI using the proton magnetic resonance spectroscopy (1H-MRS) and to see if glutamate and/or glutamine changes are associated with oxidative stress that has been reported in isolation reared animals as reviewed above. 1H-MRS is a non-invasive approach with relatively high spatial resolution and able to quantitatively measure neurochemistry in brains of living subjects [39]. It has been widely used to measure in vivo neurochemical integrity in patients with schizophrenia [29], [39] or other neuropsychiatric diseases, but few reports were available in living small animals.
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Animals and experimental procedures
Timed-pregnant Sprague Dawley rats (the Animal Center of Shantou University Medical College, Guangdong, China) were individually housed in clear polycarbonate cages (30 cm × 50 cm × 20 cm). The day of birth was designated as postnatal day 0 (PD0). On PD21 (day of weaning), twenty male rat pups from 4 litters were pooled and randomly assigned to two groups, each comprising 10 rats/group. All animal procedures described below (Fig. 1) were carried out in accordance with the guidelines laid down by the
Behavioral results
Compared to those in GH group, rats in the SI group spent less time on the open arms (86 ± 22 S vs 189 ± 29 S; p < 0.05; Fig. 3A), but more time in the closed arms (440 ± 30 S vs 302 ± 33 S; p < 0.01; Fig. 3B), indicating a higher level of anxiety. The two groups showed comparable closed arm entries (Fig. 3C), indicating that SI did not change the locomotor activity in rats.
In social interaction test, rats in the GH group stayed for a longer duration in the interaction zone around the cage with an
Discussion
In accordance with previous studies [4], [33], [49], [54], post-weaning SI for 8 weeks in the present study induced a higher level of anxiety in rats, indicated by less time spent on the open arms and more time in the closed arms of EPM. The procedure, however, had no effect on the locomotor activity of rats in the open field test performed on PD78. This result is in line with a previous report in which the locomotor activity in the open field was not modified by chronic SI in male rats [49];
Acknowledgments
This work was supported by a start-up fund from Shantou University Medical College and a postdoctoral research fellowship from Li Ka Shing foundation.
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The first two authors contributed to this work equally.