Neurosteroid [3α,5α]3‐hydroxypregnan‐20‐one inhibition of chemokine monocyte chemoattractant protein‐1 in alcohol‐preferring rat brain neurons, microglia, and astroglia

Abstract

AbstractBackgroundNeuroimmune dysfunction in alcohol use disorder (AUD) is associated with activation of myeloid differentiation primary response 88 (MyD88)‐dependent Toll‐like receptors (TLR) resulting in overexpression of the chemokine monocyte chemoattractant protein‐1 (MCP‐1/CCL2). MCP‐1 overexpression in the brain is linked to anxiety, higher alcohol intake, neuronal death, and activation of microglia observed in AUD. The neurosteroid [3α,5α][3‐hydroxypregnan‐20‐one (3α,5α‐THP) has been reported as an inhibitor of MyD88‐dependent TLR activation and MCP‐1 overexpression in mouse and human macrophages and the brain of alcohol‐preferring (P) rats.MethodsWe investigated how 3α,5α‐THP regulates MCP‐1 expression at the cellular level in P rat nucleus accumbens (NAc) and central amygdala (CeA). We focused on neurons, microglia, and astrocytes, examining the individual voxel density of MCP‐1, neuronal marker NeuN, microglial marker IBA1, astrocytic marker GFAP, and their shared voxel density, defined as intersection. Ethanol‐naïve male and female P rats were perfused 1 h after IP injections of 15 mg/kg of 3α,5α‐THP, or vehicle. The NAc and CeA were imaged using confocal microscopy following double‐immunofluorescence staining for MCP‐1 with NeuN, IBA1, and GFAP, respectively.ResultsMCP‐1 intersected with NeuN predominantly and IBA1/GFAP negligibly. 3α,5α‐THP reduced MCP‐1 expression in NeuN‐labeled cells by 38.27 ± 28.09% in male and 56.11 ± 21.46% in female NAc, also 37.99 ± 19.53% in male and 54.96 ± 30.58% in female CeA. In females, 3α,5α‐THP reduced the MCP‐1 within IBA1 and GFAP‐labeled voxels in the NAc and CeA. Conversely, in males, 3α,5α‐THP did not significantly alter the MCP‐1 within IBA1 in NAc or with GFAP in the CeA. Furthermore, 3α,5α‐THP decreased levels of IBA1 in both regions and sexes with no impact on GFAP or NeuN levels. Secondary analysis performed on data normalized to % control values indicated that no significant sex differences were present.ConclusionsThese data suggest that 3α,5α‐THP inhibits neuronal MCP‐1 expression and decreases the proliferation of microglia in P rats. These results increase our understanding of potential mechanisms for 3α,5α‐THP modulation of ethanol consumption.