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Chronic psychosocial isolation (CPSI) is known to cause several maladaptive changes in the limbic brain structures, which regulate the hypothalamic–pituitary–adrenal (HPA) axis activity. In this study, we focused our investigation on CPSI effects in the hypothalamus (HT) since it is a major driver of HPA axis activity. We also investigated whether the exposure to CPSI could alter the response to subsequent acute stress (30-min immobilization). In the HT, we followed cytosolic and nuclear levels of the glucocorticoid receptor (GR), as a mediator of HPA axis feedback inhibition, and its chaperones, the heat shock proteins (HSPs), hsp70 and hsp90. The CPSI did not cause any changes in either GR or HSPs levels. However, we observed increase of the GR and hsp70 in both HT cellular compartments as a response of naïve rats to acute stress, whereas the response of CPSI rats to acute stress was associated …