Neonatal Rat Primary Microglia: Isolation, Culturing, and Selected Applications

Abstract

Microglial cells elaborate trophic factors and cytokines and remove toxins and debris from the extracellular space, acting analogously to peripheral macrophages. Over the past two decades increased attention has been directed at the role of microglia, not only in normal physiology, but also in mediating neurotoxicity. Activation of microglia is inherent to multiple neurodegenerative disorders and exposure to toxic compounds. In large measure, these revelations have come about as a result of technologies that enable researchers to obtain high-yield and -purity primary cultures of rodent microglia. The mechanical isolation protocol discussed in this protocol offers an economical method to isolate large amount of microglia in a short and not too labor intensive manner. Most importantly, it assures a high yield of cells with great reproducibility. Given the ever increasing importance of microglia to the field of neurotoxicology research, the ability to isolate high yield of primary microglia makes it possible to investigate the role and mechanisms associated with microglial modulation of neurotoxicity. We provide a detailed description on the methods that is routinely used in our laboratory for the isolation and culturing of microglia, with emphasis on the steps which are deemed most critical for obtaining pure and healthy cultures.

Developmental Onset of Bilirubin-induced Neurotoxicity Involves Toll-like Receptor 2-dependent Signaling in Humanized UDP-glucuronosyltransferase1 Mice

“…Isolation of Primary Astrocytes and Microglia Cells from Brain-The isolation of primary glial cells was performed with modification as described previously (20). In brief, 1-3-day-old mice were decapitated and the forebrain was placed in the cold DMEM buffer (Invitrogen).…”

HAMI 3379, a CysLT2 Receptor Antagonist, Attenuates Ischemia-Like Neuronal Injury by Inhibiting Microglial Activation

“…Primary microglial cells were prepared as described previously (Candelario-Jalil et al, 2007;Ni and Aschner, 2010). In brief, the cerebral cortices were digested with 0.25% trypsin for 10 minutes at 37°C.…”

Co‐Culture of Neurons and Microglia

“…The methods described here and in Basic Protocol 3 are largely based, with modifications, on previously published protocols (Ni and Aschner, 2010; Witting and Möller, 2011). This protocol describes the first of two steps necessary to obtain primary microglia, i.e.…”