Age-associated suppression of exploratory activity during sickness is linked to meningeal lymphatic dysfunction and microglia activation [aged]

The objective of this study is to assess the contribution of meningeal lymphatic vessels to microglia response to peripheral inflammation. Aged mice (over 2 years old) were selected as a physiologically and clinically relevant model for lymphatic dysfunction because previous reporting demonstrates meningeal lymphatic function declines with age. We sequence CD11b+ cells sorted from whole brain of aged mice two hours following 1µg IL-1ß or saline vehicle intraperitoneal injection. To facilitate comparison and integration with adult mouse and experimental lymphatic ablation data, identical methods were employed. Overall design: Mice were maintained under specific pathogen-free conditions, with controlled temperature and humidity, on 12 h light:dark cycles, and ad libitum access to food and water. Experiments were all performed during light cycle. Male C57BL/6J mice over 2 years old were injected i.p. with either 1µg IL-1ß or saline vehicle (5 mice per group, 2 groups +/-IL-1ß). Two hours later, mice were euthanized by lethal dose of anesthetic (Euthasol) and perfused with cold PBS. One whole hemisphere of brain was collected from each mouse and biological replicates were pooled. Brain tissue was dissociated in 5mL Hank's buffering saline solution with DNaseI (50U/mL) and papain (4U/mL) at 37ºC for 45 minutes. Cell suspension was then passed through a 70um strainer and CD11b+ cells were magnetically sorted using CD11b MicroBeads, human and mouse (Miltenyi) as per manufacturer's instructions using AutoMACS (Miltenyi) and 2000 cells per condition were targeted for sequencing.