Dynamic association of APLNR with astrocytic remodeling in the cuprizone-treated corpus callosum

Cuprizone-induced demyelination provides a valuable experimental model for investigating glial responses and endogenous myelin repair in the central nervous system. This study examined the regional behavior of GFAP- and CD44-positive glial populations and their spatial relationship with the apelin receptor (APLNR) in the corpus callosum (CC) of 8-week-old male C57BL/6 mice. Animals were assigned to control, demyelination, and remyelination groups (n = 10/group).

Demyelination was induced by administration of 0.2% cuprizone in drinking water for 5 weeks, followed by a 5-week recovery period. Immunohistochemistry, immunofluorescence, and confocal microscopy were used to assess GFAP, CD44, APLNR, myelin basic protein (MBP), and GST-π-positive mature oligodendrocytes in the lateral (CC-L) and medial (CC-M) CC. Cuprizone induced marked myelin loss and depletion of GST-π-positive oligodendrocytes, followed by partial recovery after withdrawal.

Consistent with the established vulnerability of the rostral CC, MBP loss was greater in CC-L than in CC-M. GFAP-positive astrocytic reactivity increased markedly during demyelination and remained more persistent in CC-L during recovery, whereas CD44 showed a distinct phase-dependent regional distribution. APLNR immunoreactivity was minimal under control conditions, increased markedly during demyelination, and remained elevated during remyelination.

Its spatial overlap with both GFAP and CD44 was greatest during active demyelination, particularly in CC-L, while correlation analyses further demonstrated region- and phase-dependent relationships between APLNR and astrocytic markers. These findings demonstrate a spatially heterogeneous glial response to cuprizone-induced demyelination and identify a temporally and regionally organized association between APLNR expression and reactive astrocytic populations. The functional significance of this association remains to be established.

The present work is part of the PhD thesis of Lyubomir Gaydarski developed at the Medical University of Sofia. This study is financed by the European Union-NextGenerationEU, through the National Recovery and Resilience Plan of the Republic of Bulgaria, project BG-RRP-2.004-0004-C01 Strategic research and innovation program for development of Medical university—Sofia. Department of Anatomy, Histology and Embryology, Medical University of Sofia, 1431, Sofia, Bulgaria Lyubomir Gaydarski, Kristina Petrova, Nikola Stamenov, Alexandar Iliev, Stancho Stanchev & Boycho Landzhov Institute of Biology and Immunology of Reproduction, Bulgarian Academy of Sciences, 1113, Sofia, Bulgaria Pavel Rashev, Despina Pupaki & Milena Mourdjeva Department of Pharmacology, Pharmacotherapy and Toxicology, Medical University of Sofia, 1000, Sofia, Bulgaria Correspondence to Lyubomir Gaydarski or Boycho Landzhov.

This study was approved by the Medical-Legal Office, the Local Ethics Committee, and the Institutional Review Board of the Medical University of Sofia. All experimental procedures involving animals were conducted in accordance with Directive 2010/63/EU on the protection of animals used for scientific purposes and were approved by the Bulgarian Food Safety Agency (BFSA) under Approval Protocol No. 416/19.12.2024. The authors declare no competing interests.