IN-SITU HYBRIDIZATION FOR VASOPRESSIN MESSENGER-RNA IN THE HUMAN SUPRAOPTIC AND PARAVENTRICULAR NUCLEUS - QUANTITATIVE ASPECTS OF FORMALIN-FIXED PARAFFIN-EMBEDDED TISSUE-SECTIONS AS COMPARED TO CRYOSTAT SECTIONS

In order to study the suitability of formalin-fixed paraffin-embedded brain tissue for vasopressin (AVP)-mRNA detection, we used symmetric h alves of 5 human hypothalami. In every case, one half was formalin fix ed for 10-35 days and paraffin embedded while the other half was froze n rapidly. Following in situ hybridization (ISH) histochemistry on sys tematically obtained sections of the supraoptic (SON) and paraventricu lar nucleus (PVN) of both halves, total amounts of AVP-mRNA in these n uclei were estimated using densitometry of film autoradiographs. Total amounts of radioactivity were found to vary considerably between pati ents and amounted to 1297 +/- 302 arbitrary units (AU) (PVN) (mean +/- SEM) and 2539 +/- 346 (SON) for the cryostat sections and 868 +/- 94 (PVN) and 1259 +/- 126 (SON) for the paraffin tissue. Variations intro duced by the method itself yielded a coefficient of variation of only 0.19. Furthermore, a non-significant negative trend with postmortem de lay was found in cryostat tissue, but not in paraffin sections. No eff ect of fixation time was observed in the paraffin tissue. Both ways of tissue treatment have specific advantages and disadvantages that may be different for other probes or other brain areas. For ISH of a highl y abundant mRNA like AVP in a very heterogeneous brain area such as th e human hypothalamus, formalin-fixed paraffin-embedded tissue sections can be used for quantitative analysis of entire brain nuclei because of the small variation in this tissue, the remarkably good signal reco very (some 75% as compared to cryostat sections) and its practical adv antages with regards to anatomical orientation, storage and sampling o f the tissue.