Ventral and dorsal horn acetylcholinesterase neurons are maintained in organotypic cultures of postnatal rat spinal cord explants

Abstract Transverse sections of postnatal rat spinal cord have been cultured using the organotypic roller tube method. These explant cultures retain identifiable anatomical landmarks, allow identification of individual neurons, can be maintained for up to 8 weeks, and undergo maturational changes in vitro. Putative ventral horn motoneurons were identified in these cultures by localization to ventral horn regions analogous to those of motoneurons in vivo and by staining for choline acetyltransferase (ChAT) immunoreactivity and acetylcholinesterase (AChE) activity. Morphometric studies of the photomicrographic areas of cell bodies of these ventralhorn neurons in intact cultures show a range of sizes up to 1635 μm 2 with the average size being245 ± 7 μm 2 ( n = 724) (average ± S.E.M.). The size ranges are roughly comparable to cross-sectional areas determined previously for ventral horn motoneurons in vivo. Dorsal horn regions of these cultures also development AChE activity that was absent at explantation. Biochemical analysis of ChAT and AChE activity in pooled samples of whole cultures showed ChAT activity to be0.48 ± 0.08 ( n = 7) μmol/min/g protein and AChE activity to be2.2 ± 2.0 ( n = 7) μmol/min/g protein at 37°C (averages ± S.E.M.). These values are comparable to previously reported values for neonatal rat spinal cord in situ. Organotypic roller tube cultures of postnatal rat spinal cord provide an attractive system for studies of survival, morphology, growth and differentiation of mammalian ventral horn neurons in vitro.