Broxuridine

Social thermoregulation behaviors such as huddling among individuals can be important for energy conservation and thermoregulatory processes. Beyond that, whether huddling behavior regulates neural plasticity in the brain remains unknown. We hypothesized that huddling regulates adult neurogenesis in brain regions related to social behavior and thermoregulation. We found that cold-exposed voles had decreased aggression and increased social interaction in association with higher oxytocin but lower vasopressin hormones compared to warm-exposed voles. Cold decreased the level of cell proliferation labeling indicated by BrdU (a marker for cell proliferation) in the anterior part of the subventricular zone (SVZ) in the brain, and notably, cold-huddling (CH) voles had a higher number of proliferating cells in the hypothalamus than cold-separated (CS) voles. Moreover, CH voles displayed higher cell survival in the central amygdala and paraventricular nuclei (PVN) of the hypothalamus (both regions were related to social behavior) in comparison to CS or warm-huddling (WH) voles, respectively. Furthermore, the CH voles had more BrdU/NeuN (markers for new neurons) double-labeled cells in the SVZ than WH voles, and also more BrdU/GFAP (markers for new glial cells) double-labeled cells in SVZ and dentate gyrus compared to WH and CS voles. In addition, the newly-generated neurons differentiated into more oxytocinergic neurons in the PVN of the CH voles. Together, these data support the notion that huddling behavior is beneficial for brain plasticity by protecting cell proliferation, survival, and differentiation, and may be involved in regulating social behavior in small mammals in cold environments.

The discovery of copper(I)-catalyzed azide-alkyne cycloaddition (click chemistry) has significantly advanced the detection of proliferating cells by utilizing 5-ethynyl-2′-deoxyuridine (EdU). EdU, a thymidine analogue, is incorporated into DNA during replication and detected by the direct reaction with an azide-conjugated fluorophore. Traditionally, dividing cells are labeled using 5-bromodeoxyuridine (BrdU), another nucleotide analogue. However, BrdU detection is a harsh method that requires substantial sample processing, unlike EdU detection. EdU is classically used to identify proliferating cells; however, we report a streamlined methodology that uses EdU to label and track leukocyte recruitment that is compatible with flow cytometry and microscopy and preserves transgenic fluorophores. EdU labeling was performed in two different models of sterile inflammation: ischemic stroke and hydrochloric acid aspiration. EdU injection was timed to differentially label circulating monocytes, neutrophils and T cells. Tissue analysis showed EdU-positive monocytes and T cells were enriched in both inflammatory models. This suggests that recently divided monocytes and T cells are preferentially recruited to these vascular beds during inflammation and highlights the utility of this labeling approach to track leukocyte subtypes longitudinally during inflammation.