The discovery of carbon dioxide fixation in mammalian tissues.
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OBJECTIVE: Placental insufficiency has been considered the cause of increased morbidity in infants delivered postmaturely. Former quantitative studies have indicated a decrease in some placental structures just before term. In this study we describe a method of dual perfusion fixation to provide tissue for stereologic examination. Postmature placentas were examined with this method. STUDY DESIGN: Eleven postmature placentas and 14 placentas delivered at term were fixed by dual perfusion. The volume and the surface area of villi, the trophoblast volume, and the volume, surface area, and length of villous capillaries were estimated by stereologic examination. The Mann-Whitney test (p < or = 0.05) was used for statistical analysis. RESULTS: Morphologic features were normal in all placentas. No significant differences were disclosed in the stereologic estimates of placentas delivered at term and postmature placentas. CONCLUSION: No morphologic or significant quantitative changes were found in postmature placentas.
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A study of the distribution of phenylethanolamine-N-methyl-transferase (PNMT) activity in normal human brain is presented. After a preliminary dissection to separate brain tissue for formalin fixation and tissue designed for biochemical studies, the hemi-brain stem is cut in slices by hand and a cerebral hemisphere is cut on a cyromicrotome. "Punches" are made with an operating microscope. The dissection method was used to study the distribution of PNMT activity in 117 "punches" made on 21 slices obtained from 5 normal human brains. The caudo-rostral distribution of PNMT activity in C1 and C2 groups was found to be identical in each brain. The distribution of PNMT activity was found to be similar to that in the rat, but, in addition, important activity was found in the substantia nigra, internal pallidium and nucleus accumbens.
A method employing vascular perfusion for improved preservation of biological ultrastructure is described, and its effectiveness demonstrated for mammalian nervous tissues. Following a physiological saline flush into the aorta, hydrogen peroxide and glutaraldehyde in phosphate buffer are perfused. After buffer rinses, tissue blocks are postfixed in osmic acid and potassium ferrocyanide. The success rate is enhanced greatly by close attention to details of perfusion technique. Advantages of the method include more uniform and complete preservation. In particular, superior images of membranous elements, glycogen granules and basal laminar material are achieved. Adjustments in osmolality may render the procedure suitable for nonmammalian forms and other tissues.
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Benzodiazepine receptors can be visualized in regions of synaptic contact by electron microscopic autoradiography using [3H]flunitrazepam as a photoaffinity label in fresh brain tissue. Perfusion fixation of the tissue prior to photoaffinity labeling left the ligand binding characteristics and the light and electron microscopic distribution of benzodiazepine receptors unaltered. Therefore, the immunocytochemical localization of a neuronal marker in fixed tissue could be combined with photoaffinity labeling in order to identify the types of synapses containing benzodiazepine receptors. By using antiserum to glutamate decarboxylase, a marker of gamma-aminobutyrergic neurons, one-third of the photolabeled benzodiazepine receptors were found to be associated with immunocytochemically stained nerve endings. Thus, these synapses are the site of at lest some benzodiazepine receptors. The enhancement of gamma-aminobutyrergic synaptic transmission by benzodiazepines, shown electrophysiologically, appears to be a primary mechanism of action of this group of drugs.
Microwave devices have been developed for rapidly inactivating brain enzymes by focusing the power output into the heads of small laboratory animals. The rapid inactivation achieved prevents postmortem changes and permits the measurement of neurochemicals such as acetylcholine at concentrations close to those obtained in vivo. The technique promises the assay of neurochemical parameters not possible before.
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