In vivo labeling of cytoplasmic RNA in neurons of the immature brain cortex

Abstract

only the total and the nuclear RNAs were analyzed. The present report describes a procedure for the isolation of the neuronal cytoplasmic RNA and its application to the measurement of the approximate maturation time of its ribosomal RNA component in 8- and 20-day-old cerebral cortex perikarya. While this work was in progress a report describing the labeling pattern of neuronal and glial cytoplasmic RNA isolated from the whole brain of adult rats has appeared ~. Neuronal perikarya were isolated as described previously, except for the following modification: the solution employed for tissue suspension contained 7.5 ~ polyvinylpyrrolidone (w/v), 10 mM CaC12 and 4 ~ (w/v) instead of the 1 ~ bovine serum albumin previously used 1°. With this change in albumin concentration the suspension solution becomes isotonic. The yield and preservation of the perikaryal fraction and its purity were described 9,1°. In each experiment 10 rats were injected intrathecally with 10/~Ci of [5-aH]uridine (Amersham/Searle, 20 Ci/mmole). The neuronal perikarya were suspended in 3 ml of a solution consisting of 0.2 M sucrose- 0.06 M NaCI0.1 M Tris-HC1 buffer, pH 7.6, and the suspension was homogenized by means of 3 strokes in a glass-teflon loose-fitting homogenizer. The homogenate was centrifuged at 2,000 × g for 10 min and the supernatant was aspirated and stored in ice. This procedure was repeated and the supernatant pool was brought up to 0.25 ~ Macaloid (American Tansul Co., Houston, Texas), 0.01 M EDTA and 50/~g/ml of polyvinyl