Analysis of the vasopressin system and water regulation  in genetically polydipsic mice

Polydipsic mice, STR/N, which show extreme polydipsia and polyuria, were discovered in 1958. In the STR/N, urine outputs are much higher than in control mice. The possibility of an abnormal regulation of the arginine vasopressin (AVP) system, or an abnormality in the renal susceptibility to AVP, should be considered. In this study we investigated the AVP system and water regulation in STR/N. We sequenced the AVP and the AVP V2-receptor genes of the STR/N by direct sequencing. No mutation was found in either of them. AVP gene expression examined by in situ hybridization and plasma sodium in 8-wk-old STR/N was significantly lower than in control mice, whereas it was significantly higher at 20 wk. Renal sensitivity to injected AVP was attenuated in 20-wk-old STR/N. The suppression of AVP synthesis due to excessive water retention in 8-wk-old STR/N suggests that polydipsia may be the primary cause in this strain. The 20-wk-old STR/N became dehydrated with the acceleration of AVP synthesis, which might have resulted from secondary desensitization to AVP.

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Analysis of vasopressin gene expression by in situ hybridization and immunohistochemistry in semi-thin sections.

Journal of Histochemistry & Cytochemistry ◽

10.1177/36.11.3171163 ◽

1988 ◽

Vol 36 (11) ◽

pp. 1373-1378 ◽

Cited By ~ 18

Author(s):

A F Guitteny ◽

P Böhlen ◽

B Bloch

Keyword(s):

Gene Expression ◽

In Situ Hybridization ◽

Magnocellular Neurons ◽

Thin Sections ◽

Paraventricular Nuclei ◽

Reaction Intensity ◽

The Difference ◽

Vasopressin Gene ◽

Avp Gene

We have designed a procedure to investigate vasopressin (AVP) gene expression on plastic-embedded tissue by using in situ hybridization to detect AVP mRNA and immunohistochemistry to detect AVP. Rat brain was fixed and vibratome slices were incubated with a 45-base synthetic oligonucleotide complementary to AVP mRNA labeled with 35S, embedded in Araldite, and cut into semi-thin serial sections that were either processed for autoradiography or treated with an AVP antiserum. The results show that AVP mRNA is detectable in magnocellular neurons of the supraoptic and paraventricular nuclei in both vibratome and semi-thin sections. Osmication after hybridization does not modify the signal. AVP mRNA is restricted to the cytoplasm of magnocellular neurons and to the proximal portion of certain processes. Neurons labeled with the AVP probe were also stained with the AVP antiserum. AVP mRNA quantity and the intensity of AVP immunoreactivity are not consistently related in neurons. At least two hypotheses must be considered to explain these differences: first, the procedure presently used could lead to a reaction intensity that does not exactly reflect the amount of antigen or mRNA present in cells; second, the difference observed may reflect the fact that transcriptional and translational events are not constantly linked and can be regulated differently from one AVP neuron to another. This method provides a way to detect mRNA on semi-thin sections together with antigenic molecules and to accurately investigate gene expression in complex tissues with optimal histological quality.

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Effects of acute hypotensive stimuli on arginine vasopressin gene transcription in the rat hypothalamus

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.2000.279.4.e886 ◽

2000 ◽

Vol 279 (4) ◽

pp. E886-E892 ◽

Cited By ~ 17

Author(s):

Satoshi Kakiya ◽

Hiroshi Arima ◽

Hisashi Yokoi ◽

Takashi Murase ◽

Yuko Yambe ◽

...

Keyword(s):

In Situ Hybridization ◽

Mean Arterial Pressure ◽

Arterial Pressure ◽

Gene Transcription ◽

Arginine Vasopressin ◽

The Other ◽

Paraventricular Nuclei ◽

Vasopressin Gene ◽

Avp Gene

We investigated the baroregulation of arginine vasopressin (AVP) gene transcription in the supraoptic (SON) and paraventricular nuclei (PVN) in conscious rats by use of intronic in situ hybridization. Hemorrhage of 16 ml/kg body wt decreased mean arterial pressure (MAP) by 57% and increased both plasma AVP (control, 1.2 ± 0.3 pg/ml; 16 ml/kg body wt, 38.9 ± 3.2 pg/ml) at 10 min and AVP heteronuclear (hn)RNA levels (SON, 150%; PVN, 140% of control values) at 20 min. On the other hand, hemorrhage of 7 ml/kg body wt had no significant effect on MAP, plasma AVP, or the AVP hnRNA levels. To better understand the baroregulation, we also examined the effects of sodium nitroprusside (SNP), which induces hypotension without a change in blood volume. The subcutaneous injection of 2 mg/kg body wt SNP, which decreased the MAP by 60%, increased both plasma AVP (control, 1.6 ± 0.4 pg/ml; 2 mg/kg body wt, 8.1 ± 0.4 pg/ml) at 10 min and AVP hnRNA levels (SON, 150%; PVN, 140% of control values) at 30 min. The injection of 0.1 mg/kg body wt SNP, which reduced the MAP by 10%, failed to increase either the plasma AVP or AVP hnRNA levels. These results indicate that AVP gene transcription increases rapidly after both hypotensive hemorrhage and normovolemic hypotension. In addition, it is suggested that the set point for AVP synthesis in the baroregulation is similar to that for AVP release.

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In situ hybridization of arginine vasopressin (AVP) heteronuclear ribonucleic acid reveals increased AVP gene transcription in the rat hypothalamic paraventricular nucleus in response to emotional stress

Acta Endocrinologica ◽

10.1530/acta.0.1280466 ◽

1993 ◽

Vol 128 (5) ◽

pp. 466-472 ◽

Cited By ~ 20

Author(s):

Anne Priou ◽

Charles Oliver ◽

Michel Grino

Keyword(s):

In Situ Hybridization ◽

Gene Transcription ◽

Emotional Stress ◽

Paraventricular Nucleus ◽

Arginine Vasopressin ◽

Ribonucleic Acid ◽

Adrenal Axis ◽

Pituitary Adrenal Axis ◽

Avp Gene

The regulation of anterior pituitary adrenocorticotropin hormone (ACTH) secretion during stress involves several hypothalamic neurohormones, including arginine vasopressin (AVP). In situ hybridization techniques have been used to study the regulation of neuropeptide messenger ribonucleic acids in the hypothalamus. Owing to the relatively slow time course of the changes in cytoplasmic messenger ribonucleic acid concentrations, rapid alterations in the level of neuropeptide gene transcription could not be detected. Because of its rapid processing, the nuclear level of the heteronuclear ribonucleic acid should reflect the rate of its synthesis, namely the transcription of the gene. We have used in situ hybridization with a probe complementary to a portion of an intronic sequence of the rat AVP gene to study rapid changes in the level of AVP gene transcription during emotional stress. The specificity of our technique was demonstrated by the localization of the hybridization signals in the paraventricular nucleus, the supraoptic nucleus and the suprachiasmatic nucleus, and was confirmed by the nuclear localization of the labeling. Isolation and exposure of male rats to a novel environment induced an activation of the pituitary-adrenal axis and an increase in AVP heteronuclear ribonucleic acid concentrations in the paraventricular nucleus 2 h after the onset of the stress, suggesting that an increased AVP gene transcription may play a role in the activation of the pituitary-adrenal axis in response to emotional stress.

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RNA In Situ Hybridization for Detecting Gene Expression Patterns in the Abdomens and Wings of Drosophila Species

Methods and Protocols ◽

10.3390/mps4010020 ◽

2021 ◽

Vol 4 (1) ◽

pp. 20

Author(s):

Mujeeb Shittu ◽

Tessa Steenwinkel ◽

William Dion ◽

Nathan Ostlund ◽

Komal Raja ◽

...

Keyword(s):

Gene Expression ◽

In Situ Hybridization ◽

Expression Patterns ◽

Drosophila Species ◽

Gene Expression Patterns ◽

Probe Design ◽

Tissue Preparation ◽

Design And Synthesis ◽

Rna In Situ Hybridization

RNA in situ hybridization (ISH) is used to visualize spatio-temporal gene expression patterns with broad applications in biology and biomedicine. Here we provide a protocol for mRNA ISH in developing pupal wings and abdomens for model and non-model Drosophila species. We describe best practices in pupal staging, tissue preparation, probe design and synthesis, imaging of gene expression patterns, and image-editing techniques. This protocol has been successfully used to investigate the roles of genes underlying the evolution of novel color patterns in non-model Drosophila species.

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