Developmental regulation of epithelial sodium channel subunit mRNA expression in rat colon and lung

1998 ◽  
Vol 275 (6) ◽  
pp. G1227-G1235 ◽  
Author(s):  
Shigeru Watanabe ◽  
Kazumichi Matsushita ◽  
John B. Stokes ◽  
Paul B. McCray
Keyword(s):  
In Situ Hybridization ◽  
Epithelial Cells ◽  
Mrna Expression ◽  
Developmental Regulation ◽  
Rat Colon ◽  
Rnase Protection ◽  
Quantitative Expression ◽  
Subunit Mrna ◽  
Lung And Kidney

Na+absorption via amiloride-sensitive Na+ channels is of critical importance in the transition between fetal and neonatal life in several tissues, including the colon, lung, and kidney. To characterize and contrast the mRNA expression of each of the three epithelial Na+ channel complex (ENaC) subunits, we conducted RNase protection assays (RPA) and in situ hybridization in colon and lung in fetal (17, 19, 20, and 21 days) and postnatal (1, 3, 9, 15, and 30 days) rats (r). In the colon the α-, β-, and γ-rENaC subunits showed quantitatively different but qualitatively similar expression. All three subunits gradually increased in abundance from fetal day 19 through day 30 of life. The amount of each subunit on day 30 was approximately three times the amount at day 1. In situ hybridization showed that each subunit was localized to the surface epithelial cells with minimal expression in the crypts. The lung showed a completely different pattern. In contrast to the colon, the total amount of α-rENaC mRNA (by RPA) in the lung increased dramatically from fetal day 19 to 21, whereas β- and γ-rENaC showed modest prenatal increases. The amounts of all three mRNAs fell after birth through day 9 (to about 75% of the day 1 value). On days 15 and 30 the amount of mRNA rose to approach the values on day 1. α-rENaC mRNA abundance always exceeded β- and γ-rENaC, and the quantitative expression was different for α- than for β- and γ-rENaC. In situ hybridization studies showed that all three subunits were expressed in epithelial cells of the bronchi, bronchioles, and alveoli and not in blood vessels. These studies show striking developmental heterogeneity in rENaC mRNA expression between lung and colon, probably reflecting different developmental regulatory mechanisms in these organs.

Peptide-regulated guanylate cyclase pathways in rat colon: in situ localization of GCA, GCC, and guanylin mRNA

1993 ◽  
Vol 265 (2) ◽  
pp. G394-G402 ◽  
Author(s):  
Z. Li ◽  
M. F. Goy
Keyword(s):  
In Situ Hybridization ◽  
Mrna Expression ◽  
Natriuretic Peptide ◽  
Proximal Colon ◽  
Bacterial Toxin ◽  
Surface Epithelium ◽  
Rat Colon ◽  
Hybridization Technique ◽  
In Cells

Guanylate cyclases play a role in both physiological and pathological secretion in the mammalian intestine. Agents that raise guanosine 3',5'-cyclic monophosphate (cGMP) levels, such as atrial natriuretic peptide (ANP), guanylin (an endogenous intestinal peptide), or Escherichia coli heat-stable enterotoxin type a (STa; a bacterial toxin), enhance electrolyte secretion and the accumulation of luminal fluid. Although secretion in all parts of intestine is sensitive to changes in cGMP metabolism, an increasing body of evidence suggests that these responses are particularly important in proximal colon. To date, three peptide-sensitive membrane-bound guanylate cyclases [types A, B, and C (GCA, GCB, and GCC, respectively)] have been cloned from mammalian tissues. GCA responds to ANP, GCB to C-type natriuretic peptide, and GCC to guanylin and STa. Expression of these receptor/cyclase genes has not previously been investigated at the cellular level in the colon. Nucleotide probes specific for GCA, GCB, GCC, and guanylin were generated by polymerase chain reaction. These probes were used to evaluate colonic cyclase and guanylin mRNA expression in the rat. GCB mRNA is not detectable in this tissue either by in situ hybridization or by Northern blot analysis. In contrast, GCA, GCC, and guanylin mRNAs are all conspicuously expressed. With the in situ hybridization technique, GCA mRNA expression is seen in cells in the lamina propria. GCC mRNA expression is seen in epithelial cells throughout colonic crypts, and also, although at a slightly lower level, in cells of the surface epithelium.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression and localization of aquaporins in rat gastrointestinal tract

1999 ◽  
Vol 276 (3) ◽  
pp. C621-C627 ◽  
Author(s):  
Yu Koyama ◽  
Tadashi Yamamoto ◽  
Tatsuo Tani ◽  
Kouei Nihei ◽  
Daisuke Kondo ◽  
...  
Keyword(s):  
Small Intestine ◽  
In Situ Hybridization ◽  
Gastrointestinal Tract ◽  
Epithelial Cells ◽  
Cellular Localization ◽  
Basolateral Membrane ◽  
Rnase Protection Assay ◽  
Rnase Protection ◽  
Basolateral Membranes

A family of water-selective channels, aquaporins (AQP), has been demonstrated in various organs and tissues. However, the localization and expression of the AQP family members in the gastrointestinal tract have not been entirely elucidated. This study aimed to demonstrate the expression and distribution of several types of the AQP family and to speculate on their role in water transport in the rat gastrointestinal tract. By RNase protection assay, expression of AQP1–5 and AQP8 was examined in various portions through the gastrointestinal tract. AQP1 and AQP3 mRNAs were diffusely expressed from esophagus to colon, and their expression was relatively intense in the small intestine and colon. In contrast, AQP4 mRNA was selectively expressed in the stomach and small intestine and AQP8 mRNA in the jejunum and colon. Immunohistochemistry and in situ hybridization demonstrated cellular localization of these AQP in these portions. AQP1 was localized on endothelial cells of lymphatic vessels in the submucosa and lamina propria throughout the gastrointestinal tract. AQP3 was detected on the circumferential plasma membranes of stratified squamous epithelial cells in the esophagus and basolateral membranes of cardiac gland epithelia in the lower stomach and of surface columnar epithelia in the colon. However, AQP3 was not apparently detected in the small intestine. AQP4 was present on the basolateral membrane of the parietal cells in the lower stomach and selectively in the basolateral membranes of deep intestinal gland cells in the small intestine. AQP8 mRNA expression was demonstrated in the absorptive columnar epithelial cells of the jejunum and colon by in situ hybridization. These findings may indicate that water crosses the epithelial layer through these water channels, suggesting a possible role of the transcellular route for water intake or outlet in the gastrointestinal tract.

In situ hybridization histochemistry as a method to assess GABAAreceptor subunit mRNA expression following chronic alprazolam administration

1999 ◽  
Vol 13 (3) ◽  
pp. 211-218 ◽  
Author(s):  
Jeanne M. Fahey ◽  
Gary A. Pritchard ◽  
Jeffrey M. Grassi ◽  
John S. Pratt ◽  
Richard I. Shader ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Mrna Expression ◽  
Hybridization Histochemistry ◽  
Subunit Mrna ◽  
In Situ Hybridization Histochemistry

scholarly journals Temporal changes in inhibin subunit mRNAs during atresia of preovulatory follicles in the rat

1998 ◽  
Vol 159 (2) ◽  
pp. 331-340 ◽  
Author(s):  
JT Uilenbroek ◽  
AL Durlinger ◽  
M Tebar ◽  
P Kramer ◽  
RH van Schaik ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Gnrh Antagonist ◽  
Rnase Protection Assay ◽  
Protection Assay ◽  
Rnase Protection ◽  
Subunit Mrna ◽  
Preovulatory Follicles ◽  
Inhibin Subunits

This study aimed to investigate the time course of disappearance of the mRNAs of the various subunits of inhibin in follicles which become atretic. An animal model was used in which atresia of preovulatory follicles could be studied in a chronological order. Injection of gonadotrophin-releasing hormone (GnRH) antagonist (20 microg) at the morning of pro-oestrus (P) blocked ovulation and the 10-12 preovulatory follicles became gradually atretic. A second injection was given the next day to prevent delayed ovulation. The rate of atresia could be delayed by simultaneous administration of a subovulatory dose of human chorionic gonadotrophin (hCG) (0.5 IU) and could be advanced by administration of a fivefold larger amount of GnRH antagonist. Functional activity of follicles becoming atretic was studied by measuring oestradiol production after incubation of individual follicles for 4 h. Follicles isolated 24 h after the first injection of GnRH antagonist (P+24) already secreted significantly less oestradiol in vitro than follicles isolated at pro-oestrus, although they were morphologically not different from pro-oestrous follicles. Follicles isolated at P+24 from hCG-treated rats secreted more oestradiol compared with follicles from rats not treated with hCG. In contrast, follicles isolated at P+24 from rats that were given a fivefold larger amount of GnRH antagonist secreted less oestradiol. Once this model was validated, temporal changes in inhibin subunit mRNAs in follicles undergoing atresia were measured by in situ hybridization and RNase protection assay. In situ hybridization showed abundant alpha- and betaA-subunit mRNA in the whole granulosa layer of preovulatory follicles at P and P+24, while betaB-subunit mRNA was restricted to the antral layer and cumulus. At P+48 the amount of alpha- and betaA-subunit mRNA had declined and was restricted to the cumulus, whereas betaB-subunit mRNA was absent. In the atretic follicles present at P+72 and P+96, mRNAs of all three inhibin subunits were absent. Administration of 0.5 IU hCG delayed the decline in the amount of alpha, betaA and betaB mRNA in preovulatory follicles at P+48. RNase protection assay of inhibin subunits in isolated follicles revealed no changes between P and P+24. However, at P+48, the mRNAs of alpha- and betaA-subunits were decreased. Expression of the mRNA of betaB-subunit declined gradually from P to P+48. The present study demonstrates that in follicles which are becoming atretic, mRNAs of alpha- and betaA-subunits decline simultaneously with the appearance of pycnotic cells in the granulosa layer, while betaB-subunit mRNA declines earlier, simultaneously with the decrease in the ability to secrete oestradiol in vitro.

Developmental expression of human angiotensinogen in transgenic mice

1998 ◽  
Vol 274 (5) ◽  
pp. F932-F939 ◽  
Author(s):  
Gongyu Yang ◽  
Curt D. Sigmund
Keyword(s):  
In Situ Hybridization ◽  
Transgenic Mice ◽  
Developmental Regulation ◽  
Rnase Protection Assay ◽  
Late Gestation ◽  
Developmental Expression ◽  
Rnase Protection ◽  
Adult Mice ◽  
Specific Regulation

Transgenic mice containing the human angiotensinogen ( HAGT) gene were utilized to determine the developmental regulation of HAGT expression. RNase protection assay on total RNA obtained from whole transgenic fetuses revealed that HAGT expression was first detected at embryonic day 8.5( E8.5) and was abundant from E9.5 onward. The earliest expression of the HAGT transgene appeared to precede the earliest expression of the endogenous mouse AGT gene by 1–2 days. Northern blot analysis revealed moderate levels of HAGT mRNA in liver and kidney and low levels of HAGT mRNA in heart and brain from E16.5 ( day 16.5 of gestation) onward. HAGT mRNA in liver, although abundant during late gestation and in 2-wk-old and adult mice, decreased transiently around birth. In situ hybridization performed on sections from whole fetuses revealed that HAGTmRNA was restricted to the developing liver and heart between E9.5 and E11.5 but became more widespread to include the developing aorta, brain, subcutaneous tissues, and vertebra at E13.5. In situ hybridization analysis on fetal kidneys from late gestation, newborn, and 2-wk-old mice demonstrated a progressive restriction of HAGT mRNA to developing cortical proximal tubular cells. These data illustrate the developmental tissue-specific regulation of HAGTexpression and demonstrate that sequences present in the transgene can confer an appropriate developmental expression profile.

In situ hybridization localization of the Na+ channel β1 subunit mRNA in rat CNS neurons

1994 ◽  
Vol 176 (1) ◽  
pp. 119-122 ◽  
Author(s):  
Youngsuk Oh ◽  
Shunsuke Sashihara ◽  
Stephen G. Waxman
Keyword(s):  
In Situ Hybridization ◽  
Na Channel ◽  
Subunit Mrna ◽  
Cns Neurons ◽  
Rat Cns
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