Brain-wide mapping reveals widespread GLUT4 downregulation and region-specific perineuronal net remodeling following prolonged hypercaloric diet

Metabolic disorders trigger low-grade brain inflammation, disrupting neuronal homeostasis and plasticity. A key factor is impaired glucose handling: besides insulin-independent transport, the insulin-sensitive transporter GLUT4 ensures energy supply in specific metabolic conditions, and its dysfunction promotes brain insulin resistance and cognitive decline. Energy deficits may also affect the extracellular matrix (ECM), which sustains circuit stability. Among ECM components, perineuronal nets (PNNs), structures surrounding mainly parvalbumin-positive (PV+) interneurons, are crucial for network regulation. However, there is still little information on the distribution of GLUT4 in the whole brain and the possible link with ECM plasticity players, both in normal physiological conditions and in metabolic challenge. Here, we developed a high-throughput automated workflow to map brain-wide GLUT4, PNNs, and PV+ cells in mice maintained on a standard diet (SD) or a high-fat diet (HFD) for 12 months. To evaluate whether metabolic normalization could restore impaired marker expression, two additional HFD groups were treated with widely used antidiabetic drugs: the DPP-4 inhibitor linagliptin and the sulfonylurea glimepiride. In SD-fed mice, GLUT4 expression was mainly found in the gray matter, with highest levels in the cortex. Chronic HFD induced a type 2 diabetes-like phenotype accompanied by a nearly global reduction of GLUT4 and cortex-specific PNN alterations. Pharmacological treatments produced modest effects, failing to restore central GLUT4 or PNN expression. These findings highlight that metabolic stress disrupts both GLUT4 and PNN homeostasis and suggest that preserving brain health under metabolic challenge demands therapeutic approaches that extend beyond peripheral glucose regulation.