Human primary mixed brain cultures: preparation, differentiation, characterization and application to neuroscience research

Abstract

BackgroundCulturing primary cortical neurons is an essential neuroscience technique. However, most cultures are derived from rodent brains and standard protocols for human brain cultures are sparse. Herein, we describe preparation, maintenance and major characteristics of a primary human mixed brain culture, including neurons, obtained from legally aborted fetal brain tissue. This approach employs standard materials and techniques used in the preparation of rodent neuron cultures, with critical modifications.ResultsThis culture has distinct differences from rodent cultures. Specifically, a significant numbers of cells in the human culture are derived from progenitor cells, and the yield and survival of the cells grossly depend on the presence of bFGF. In the presence of bFGF, this culture can be maintained for an extended period. Abundant productions of amyloid-β, tau and proteins make this a powerful model for Alzheimer’s research. The culture also produces glia and different sub-types of neurons.ConclusionWe provide a well-characterized methodology for human mixed brain cultures useful to test therapeutic agents under various conditions, and to carry forward mechanistic and translational studies for several brain disorders.Electronic supplementary materialThe online version of this article (doi:10.1186/s13041-014-0063-0) contains supplementary material, which is available to authorized users.

Initial analysis of peripheral lymphocytic extracellular signal related kinase activation in autism

“…These cells were generally seen has having an empty or hollow area in the cell due to lack of staining in this region. These cells were counted using the count feature in Adobe Photoshop (Ray et al, 2014). …”

“…Protein standard (BioRad #161–0374) was loaded in one lane of the gel as a marker. Human fetal neuron (HFN) lysate (Ray et al, 2014) and human brain lysate (Long et al, 2014) were also loaded as positive controls for p-ERK expression. Gel was run at 200 V for 1.5 h, and transfer was performed using the Invitrogen iBlot system.…”

Potential for Stem Cells Therapy in Alzheimer’s Disease: Do Neurotrophic Factors Play Critical Role?

“…Abundant tau and amyloid-β production in human brain cultures provides a powerful cellular model for AD. In a recent study Ray et al provide a well-characterized methodology for fetal human primary brain cell culture, which is useful to test the therapeutic efficacy of drugs targeting AD [28]. Cultures of induced pluripotent stem cell (iPSC) generated from fibroblasts of FAD patients with presenilin 1 and presenilin 2 mutations were characterized after acquiring neuronal lineage [29].…”

Analysis of Neural Stem Cells from Human Cortical Brain Structures In Vitro

“…Some papers presented morphological and molecular characteristics of NSPC derived from the forebrain of human fetuses on gestation weeks 7-12, which corresponds to the period of active cortical neurogenesis [3,4,31,33], while generation of neurons in human neocortex during the second trimester decreases [25]. Morphological studies showed that this period is characterized by migration and differentiation of neurons, increase in cell mass, and structural changes [8,25] determining specifi c properties of human brain cortex that distinguish it from the brain cortex of animals [35].…”

“…In humans, active neurogenesis in the neocortex lasts 65 days, from day 50 (week 7) through day 115 (week 17) of fetal development [2] and signifi cantly decreased by gestation weeks 18-24 [25,35]. NSPC populations in humans and rodents differ by cell composition [16] during in vivo and in vitro differentiation [7,13,17,24,33]. All these differences indicate the necessity of using human fetal neural cells as the positive control during the development of advanced methods of clinical cell therapy of neural disorders in humans based on embryonic or induced pluripotent stem cells [27].…”