4 " ," r" ELSEVIER Molecular Brain Research 28 (1995) 311-318 MOLECULAR BRAIN RESEARCH Research report Tissue pH as an indicator of mRNA preservation in human post-mortem brain Ann E. Kingsbury a Oliver J.F. Foster b,* Angus P. Nisbet c, Nigel Cairns d, Louise Bray c, David J. Eve c, Andrew J. Lees e, C. David Marsden b a MRC Human Movement and Balance Unit, seconded to the Parkinson's Disease Society Brain Bank, Institute of Neurology, 1 Wakefield Street, London WC1N 1P J, UK b Institute of Neurology, Queen Square, London WCIN3BG, UK c Parkinson's Disease Society Brain Bank, Institute of Neurology, 1 Wakefield Street, London WCIN 1PJ, UK d Medical Research Council Alzheimer's Disease Brain Bank, Institute of Psychiatry, De Crespigny Park, London SE5 8AF, UK e National Hospital for Neurology and Neurosurgery, Queen Square, London WCIN 3BG, UK Accepted 20 September 1994 Abstract The relationship between pH and mRNA preservation in post-mortem human brain was examined using in situ hybridization histochemistry and Northern hybridization with oligonucleotide probes in a large group of human subjects, including control and neuropathological cases. Tissue pH was found to correlate strongly with preservation of four mRNA species in three brain areas. Tissue with low pH, assumed to result from prolonged terminal hypoxia, contained reduced or absent mRNA, while tissue with higher pH was found to contain quantifiable amounts, the values for pathological brain samples being comparable to those for control material of similar pH. Measurement of tissue pH provides a simple means to screen post-mortem brain for mRNA preservation and is suggested as a means to match material in case-control studies of human neurodegenerative disease. Keywords: Human brain; Post-mortem; pH; mRNA preservation; In situ hybridization I. Introduction Although in situ hybridization histochemistry (ISHH) is a well-established technique in neurobiology it has only been applied to the study of human post-mortem brain relatively recently. The usefulness of this tech- nique in human post-mortem studies is compromised by considerable inter-individual variability, thought to result from peri-mortem changes, which confounds the results of quantitative case-control studies of gene ex- pression in human neurodegenerative disease. It is now generally agreed (for review see Barton et al. [1]) that while post-mortem delay may critically affect the preservation of, for instance, catecholamines and some peptides in the brain, many molecules, in- cluding mRNA are preserved for long periods in post- " Corresponding author. Parkinson's Disease Society Brain Bank, 1 Wakefield Street, London WC1N 1PJ, UK. Fax: (44) 71-278 4993. 0169-328X/95/$09.50 © 1995 Elsevier Science B.V. All rights reserved SSDI 0169-328X(94)00219-3 mortem tissue. In contrast, evidence has accumulated to show that ante-mortem events in the brain, particu- larly hypoxia, exert an important influence on a num- ber of neurochemical parameters, mRNA preservation [11], enzyme activities [23,19], and neurotransmitter and neuropeptide concentrations [6,13] have all been shown to vary significantly with varying agonal state. Animal studies confirm the influence of terminal hypoxia on RNA preservation [2,12] and have also demonstrated a close relationship between hypoxia and the pH both of brain tissue and CSF [3,17]. Studies of human post-mortem tissue and CSF have demon- strated a significant negative correlation between pH value and lactate concentration [23], low tissue pH being thought to reflect anaerobic glycolysis within the brain, more lactate accumulating with greater respira- tory distress or a longer terminal phase [9]. Tissue pH is therefore regarded as a useful marker for terminal hypoxia [20]. In order to elucidate the role of agonal state in