Organotypic brain slice cultures: A review

Abstract

In vitro cell cultures are an important tool for obtaining insights into cellular processes in an isolated system and a supplement to in vivo animal experiments. While primary dissociated cultures permit a single homogeneous cell population to be studied, there is a clear need to explore the function of brain cells in a three-dimensional system where the main architecture of the cells is preserved. Thus, organotypic brain slice cultures have proven to be very useful in investigating cellular and molecular processes of the brain in vitro. This review summarizes (1) the historical development of organotypic brain slices focusing on the membrane technology, (2) methodological aspects regarding culturing procedures, age of donors or media, (3) whether the cholinergic neurons serve as a model of neurodegeneration in Alzheimer's disease, (4) or the nigrostriatal dopaminergic neurons as a model of Parkinson's disease and (5) how the vascular network can be studied, especially with regard to a synthetic blood-brain barrier. This review will also highlight some limits of the model and give an outlook on future applications.

Keywords: cholinergic; dopaminergic; organotypic; vascular; whole-brain cultures.

Fig. 1

Organotypic brain slices are prepared from whole postnatal or adult brains (A), and 100- to 400-μm-thick sections are cut with a vibratome (B) and placed in an insert with 0.4-μm semipermeable pores (C). Cholinergic neurons stained for choline acetyltransferase⁺ neurons were found in the striatum, septum and basal nucleus of Meynert after incubation with 10 ng/ml nerve growth factor (NGF) for two weeks (D). Dopaminergic tyrosine hydroxylase⁺ neurons survive well in the ventral mesencephalon (vMES) when incubated with 10 ng/ml glial cell line-derived neurotrophic factor (GDNF) for two weeks (E). Immunostainings for tryptophane hydroxylase show raphe neurons after two weeks in culture (F). Brain slices display a strong vascular network, as seen by collagen IV staining (Alexa-488, green) (G). The vascular system is in direct interaction with astrocytes co-stained for collagen IV (Alexa-488, green) and glial fibrillary acidic protein (GFAP, Alexa-546, red) (H). High-power confocal microscopy shows a collagen IV⁺ (Alexa-488, green) vessel in the organotypic slices after two weeks in culture (I). Adult whole-brain organotypic slices (110-μm thick) of the APP_SweDI Alzheimer mouse model were cultured for two weeks, stained for beta-amyloid using Alexa-546 (red) (J, K, L) and co-stained with GFAP (Alexa-488, green) for reactive astrocytes (K) or collagen IV using Alexa-488 (green) (L). Note that sections in Figure D-F&J were stained using the chromogenic substance DAB, while the sections in G–L underwent fluorescent staining. Scale bar in A = 980 μm (A–C), 300 μm (D-striatum), 600 μm (D-septum), 90 μm (D-nBM); 600 μm (E upper), 20 μm (E lower), 70 μm (F), 250 μm (G), 60 μm (H), 30 μm (I), 240 μm (J), 36 μm (K) and 24 μm (L).