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F M Correa

Publications and source records attributed to F M Correa.

At least 37 records · Page 2Linked to original sources
PubMed1988

Radioimmunochemical methods for the quantitative autoradiographic determination of antigens in brain and other tissues.

1. We have used horseradish peroxidase-conjugated protein A- and 125I-protein A to develop immunohistochemical and radioimmunohistochemical methods for the localization of antigens in brain and other tissues of the rat. 2. We visualized methionine-enkephalin fibers in the rat brain by incubating tissue sections with a specific polyclonal antibody and peroxidase-conjugated protein A. The method is simple, fast, and less expensive and more sensitive than classical immunohistochemical techniques and the principle could be used to visualize many other tissue antigens. 3. Incubation of tissue samples with specific polyclonal antibodies and 125I-protein A, followed by autoradiography, allows the permanent recording of the radioimmunohistochemical localization of brain methionine-enkephalin, tyrosine hydroxylase, and angiotensin-converting enzyme and of pituitary vasopressin and could be applied to the localization of many other tissue antigens. 4. A new quantitative radioimmunohistochemical technique for methionine-enkephalin allows the determination of the endogenous peptide content in discrete brain nuclei from 16-microns-thick sections. The method is based on the quantitative determination of the amount of 125I-protein A bound to specific tissue areas after incubation with a specific polyclonal antibody, followed by autoradiography and computerized microdensitometry. To quantify the endogenous peptide content, the values obtained are interpolated into a methionine-enkephalin internal standard curve. This standard curve was constructed by measuring endogenous concentrations of methionine-enkephalin by radioimmunoassay in specific brain regions and correlating these values with quantitative autoradiographic determinations in homologous areas of adjacent sections. Similar methods can be developed for other tissue antigens. 5. These new methods allow for the localization and quantification of tissue antigens in very discrete areas of the brain and other tissues and have a wide application in neurobiology and pathology.

Animals↗
PubMed1988

Quantitative radioimmunohistochemical method using [125I]-protein A to measure the content of methionine enkephalin in discrete rat brain areas.

We report a quantitative radioimmunohistochemical method, using [125I]-protein A in combination with a specific antibody to methionine enkephalin (Met-enk), for determination of the content of this peptide in discrete areas of rat brain. After paraformaldehyde fixation, rat brain sections were incubated with a Met-enk polyclonal antibody, followed by incubation with [125I]-protein A. After autoradiography with 3H-sensitive Ultrofilm, optical densities (OD) were quantified by computerized microdensitometry. The OD obtained were compared to a standard curve, constructed after determination by radioimmunoassay of the Met-enk content in corresponding brain areas from adjacent tissue sections. After comparing 15 different brain areas over a ninetyfold range of concentrations, we found a linear relationship between the content of Met-enk, as determined by radioimmunoassay, and the OD generated by autoradiography. The content of Met-enk in other discrete brain areas can be quantified by interpolation of the OD determined by autoradiography in the standard curve. The method allows, for the first time, precise quantification of peptide concentrations in discrete areas and nuclei from thin sections of rat brain. This technique has a more than 100-fold higher sensitivity than classical radioimmunoassays, with the additional advantage of neuroanatomical localization. It also has the potential for application to the quantification of many other antigens present in brain and other tissues.

Animals↗
PubMed1986

Quantitative distribution of angiotensin-converting enzyme (kininase II) in discrete areas of the rat brain by autoradiography with computerized microdensitometry.

We report the localization of angiotensin-converting enzyme (kininase II, EC 3.4.15.1) in discrete nuclei and areas of the rat brain by a quantitative autoradiographic technique using image processing coupled to computerized microdensitometry, after incubation of brain sections with the specific converting enzyme inhibitor [125I]351A. High angiotensin-converting enzyme levels are present in circumventricular organs (organon subfornicalis and area postrema), the choroid plexus, and extrapyramidal areas (nucleus caudatus, globus pallidus and substantia nigra) with intermediate levels in selected hypothalamic, septal, habenular and brainstem nuclei. Our results support the idea that angiotensin II could be formed in specific brain areas, both outside and inside the blood-brain barrier. In other brain structures, such as the extrapyramidal areas, kininase II could be involved in the processing or metabolism of other brain peptides.

Animals↗
PubMed1986

Angiotensin II binding sites in the anteroventral-third ventricle (AV3V) area and related structures of the rat brain.

Angiotensin II binding sites were localized in the rat brain by incubation of sagittal sections with 125I-[Sar1]-angiotensin II, followed by [3H]Ultrofilm autoradiography. Binding sites were localized in the subfornical organ and the entire anteroventral-third ventricle (AV3V) area, including the subependymal area of the anterior third ventricle, the median preoptic nucleus and the organon vasculosum laminae terminalis. Binding sites were also present in the paraventricular and periventricular nuclei and in the median eminence. These findings support the hypothesis of an extensive involvement of angiotensin in the neuronal circuits connecting the subfornical organ, the AV3V area, the paraventricular nucleus and the median eminence, and in the regulation of fluid balance, blood pressure and pituitary secretion.

Animals↗
PubMed1986

Increased atrial natriuretic peptide (6-33) binding sites in the subfornical organ of water deprived and Brattleboro rats.

Binding sites for rat atrial natriuretic peptide (6-33) (ANP) were quantitated in the subfornical organ of chronically dehydrated homozygous Brattleboro rats unable to synthesize vasopressin; heterozygous Brattleboro rats, their controls, Long Evans rats and Long Evans rats after 4 days of water deprivation. Brain sections were incubated in the presence of 125I-ANP and the results analyzed by autoradiography coupled to computerized microdensitometry and comparison to 125I-standards. Brattleboro rats and water deprived Long Evans rats presented a higher number of ANP binding sites than their normally hydrated controls. Our results suggest a role of ANP binding sites in the subfornical organ in the central regulation of fluid balance and vasopressin secretion.

Animals↗
PubMed1986

Increased number of angiotensin II receptors in the subfornical organ of spontaneously hypertensive rats.

Angiotensin II receptors were studied in the brain of spontaneously hypertensive rats (SHR) and their normotensive controls, Wistar-Kyoto rats (WKY) by quantitative autoradiographic methods coupled to computerized microdensitometry and comparison with 125I-standards, after incubation of brain sections with 125I-Sar1-angiotensin II. Binding sites for angiotensin II were highly localized in the subfornical organ. The number of binding sites was higher in the subfornical organ of young (4-week-old) and adult (14-week-old) SHR when compared with WKY rats (young rats: 735 +/- 91 and 444 +/- 85 fmol/mg protein, SHR and WKY, respectively; adult rats: 1390 +/- 64 and 848 +/- 136 fmol/mg protein, SHR and WKY, respectively). Our results suggest that subfornical organ angiotensin II receptors are involved in genetic hypertension in the rat. An increased number of subfornical organ angiotensin II receptors substantiates the idea of an overactive central angiotensin II system in spontaneous hypertension.

Angiotensin II↗
PubMed1985

Quantitative autoradiographic determination of angiotensin-converting enzyme binding in rat pituitary and adrenal glands with 125I-351A, a specific inhibitor.

We describe a quantitative autoradiographic technique which allows measurement of angiotensin-I-converting enzyme [ACE] (kininase II, peptidyldipeptide hydrolase, EC 3.4.15.1) levels in discrete areas of pituitary and adrenal glands in individual animals. Tissue sections were incubated with 125I-351A, a specific ACE inhibitor, and results were obtained with computerized densitometry and comparison to 125I standards. There were high levels of ACE in both the anterior and posterior lobes of the pituitary, with no detectable binding in the intermediate lobe. The maximum binding capacity (Bmax) was 920 +/- 62 fmol/mg protein for the anterior pituitary and 1162 +/- 67 fmol/mg protein for posterior pituitary. The binding affinity constant (Ka) was 0.95 +/- 0.11 X 10(9) M-1 and 1.20 +/- 0.19 X 10(9) M-1 for the anterior and posterior lobes, respectively. In the adrenal gland, there were two distinct areas of specific binding, the adrenal medulla and the adrenal capsule-zona glomerulosa area. The Bmax for the adrenal medulla was 652 +/- 80 fmol/mg protein and 294 +/- 53 fmol/mg protein for the adrenal capsule-zona glomerulosa. The Ka for 351A was 1.04 +/- 0.19 X 10(9) M-1 and 1.74 +/- 0.40 X 10(9) M-1 for medulla and adrenal capsule-zona glomerulosa respectively. The results support the existence of local ANG systems active in both the pituitary and adrenal glands.

Adrenal Medulla↗
PubMed1985

Quantitative autoradiographic determination of angiotensin-converting enzyme (kininase II) binding in individual rat brain nuclei with 125I-351A, a specific enzyme inhibitor.

We report the localization and characterization of angiotensin-converting enzyme (kininase II) in discrete nuclei from individual rat brains by a quantitative autoradiographic technique coupled to computerized microdensitometry. The enzyme was quantitated by incubation of 16-micron-thick brain sections with 0.07-2 nM of the converting enzyme inhibitor 125I-351A and comparison to 125I-standards. This technique can be applied to the study of other enzymes in single rat brain nuclei.

Animals↗
PubMed1985

Femtomole isotopic-enzymatic microassay of histamine in microgram amounts of brain tissue.

We present a modification of a previously described radioenzymatic technique (Snyder et al., Journal of Pharmacology and Experimental Therapeutics, 153: 544-549, 1966) for the determination of histamine. This assay is based on the incubation of tissue histamine with a partially purified preparation of histamine-N-methyltransferase (S-adenosyl-L-methionine: histamine N-methyltransferase, E.C. 2.1.1.8; HMT) in the presence of the natural donor of methyl groups, [3H]-methyl-S-adenosyl-L-methionine ([3H]-SAMe). We have found that HMT retains its histamine N-methylating activity at 4 degrees C. Incubation at low temperature results in a considerable increase in the sample to blank ratio. The reduction of the total amount of [3H]-SAMe used and addition of anhydrous sodium sulfate before the last organic extraction step further reduced the blank levels. With these modifications, the sensitivity of the method was increased to the lower femtomole range. The present assay can be used for the determination of as little as 1 to 2 pg of histamine in samples from individual rat brain nuclei containing 2 to 10 micrograms of protein.

Animals↗
PubMed1984

Quantitative determination of angiotensin II binding sites in rat brain and pituitary gland by autoradiography.

Rat brain and pituitary angiotensin II (AII) binding sites were quantitated by incubation of tissue sections with 125I-[Sar1] AII, Ultrofilm radioautography, computerized densitometry, and comparison with 125I-standards at appropriate film exposure times. The highest number of AII binding sites was found in anterior pituitary and the circumventricular organs, organon subfornicalis and organon vasculosum laminae terminalis.

Angiotensin II↗
PubMed1984

Radioimmunoassay of met-enkephalin in microdissected areas of paraformaldehyde-fixed rat brain.

We studied the effect of various sample preparation procedures on rat brain met-enkephalin content, measured by radioimmunoassay. Whole brain met-enkephalin content of rats killed by decapitation followed by immediate tissue freezing was similar to that of rats killed by microwave irradiation and to those of rats anesthetized with pentobarbital or halothane before killing, whether previously perfused with paraformaldehyde or not. In contrast, a decrease (up to 80%) in met-enkephalin concentrations was observed when brain samples were frozen and thawed to mimic the procedure utilized in the "punch" technique for analysis of discrete brain nuclei. This decrease was totally prevented by paraformaldehyde perfusion of the brain prior to sacrifice. Brain perfusion did not alter the amount of immunoassayable met-enkephalin extracted from tissue or its profile after Sephadex chromatography. Paraformaldehyde perfusion results in better morphological tissue preservation and facilitates the "punch" dissecting technique. Paraformaldehyde perfusion may be the procedure of choice for the measurement of neuropeptides in specific brain nuclei dissected by the "punch" technique.

Anesthesia↗
PubMed1983

High histamine levels in specific hypothalamic nuclei of Brattleboro rats lacking vasopressin.

Histamine levels were determined in whole hypothalamus and specific hypothalamic nuclei of male 9-week-old homozygous Brattleboro rats lacking vasopressin and presenting the syndrome of diabetes insipidus (DI rats). These levels were compared to those in heterozygous Brattleboro rats with a partial deficit in vasopressin (HZ rats) and those in Long Evans control rats (LE rats). In whole hypothalamus, histamine levels in DI rats were higher than those found in HZ and LE rats. DI rats showed histamine levels higher than those present in LE rats in the nuclei supraopticus, paraventricularis and suprachiasmatis and in the area retrochiasmatica. Vasopressin replacement produced a selective decrease in histamine levels of DI rats, restricted to the area retrochiasmatica and nuclei supraopticus and paraventricularis. In contrast, vasopressin increased histamine levels in the area retrochiasmatica and eminentia mediana of HZ rats. In other areas normally rich in vasopressin such as the nucleus arcuatus and the eminentia mediana, the histamine content was not different among LE, HZ or DI rats. Our results suggest a physiological interaction between vasopressin and histamine systems in specific hypothalamic areas of the rat and support the hypothesis of a role of brain histamine on the central control of water balance.

Animals↗
PubMed1983

Somatostatin inhibits the isoproterenol-stimulated adenylate cyclase in the intermediate lobe of the male rat pituitary gland.

Somatostatin-14 inhibited the isoproterenol-stimulated adenylate cyclase in cell-free homogenates of pituitary intermediate lobe of the male rat with a Ki of 7.25 +/- 0.31 X 10(-7) M. Somatostatin-28 was found to be a partial agonist. Other neuropeptides (met- and leu-enkephalin, beta-endorphin, alpha-melanocyte-stimulating hormone, ACTH, neurotensin, substance P, vasoactive intestinal polypeptide, and vasopressin) were without effect. Our results suggest a role for pituitary and/or brain somatostatin in the physiological regulation of the intermediate pituitary lobe, and the possibility of an interaction between somatostatins and catecholamines on the regulation of intermediate lobe adenylate cyclase.

Adenylyl Cyclases↗
PubMed1981

Diffuse enkephalin innervation from caudate to globus pallidus.

The source of enkephalin in neuronal terminals of the rat globus pallidus was evaluated with electrolytic and knife cut lesions. Straight knife cuts medial to the globus pallidus or a curved knife cut just below the corpus callosum do not reduce immunofluorescence staining in the globus pallidus, indicating that cerebral cortex is not a major source of pallidal enkephalin. Electrolytic lesions of dorsal and ventral portions of the globus pallidus, and even lesions which sever medial from lateral globus, fail to reduce pallidal staining, ruling out long enkephalin neurons traversing the globus. Large electrolytic lesions of the head of the caudate do decrease dorsal pallidal fluorescence, though multiple small caudate lesions are without observable effect. Large pallidal lesions elicit a build-up of enkephalin fluorescence in the caudate. These results suggest that the caudate is the sole source of pallidal enkephalin and that the innervation diffusely converges on the smaller globus pallidus.

Animals↗