In situ hybridization of the vasopressin mRNA in the rat hypothalamus by use of a synthetic oligonucleotide probe

Nojiri, Hiroshi; Sato, Moriyuki; Urano, Akihisa

doi:10.1016/0304-3940(85)90336-2

Cited by 42 publications

(9 citation statements)

References 6 publications

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“…4C) by use of a 45-mer oligoprobe corresponding to the rat vasopressin mRNA, labeled at its 5' end with 32P. This result correlates with similar works [Lewis et al, 1985;Nojiri et al, 1985;Uhl et al, 19851 and demonstrates the potency of and interest in using synthetic oligonucleotides to identify related sequences of bases in tissues.…”

Section: Study Of Gene Expressionsupporting

confidence: 90%

In situ hybridization histochemistry for the analysis of gene expression in the endocrine and central nervous system tissues: A 3‐year experience

Bloch

Popovici

Guellec

et al. 1986

J of Neuroscience Research

130

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We report our experience in development of the in situ hybridization (ISH) procedure to detect messenger RNAs (mRNAs) coding for various molecules involved in endocrine glands and central nervous system activity, including mRNAs coding for endorphin precursors [preproenkephalin A (PPA), pro-opiocortin (POMC)], vasopressin, and transferrin. Various conditions of fixation and handling of the tissues were tested to establish optimal parameters for mRNA detection. Double-stranded DNA probes labeled by nick translation, synthetic oligonucleotides labeled at their 5' end, as well as single-stranded RNA probes were used, after incorporation of 32P- or 35S-labeled nucleotides. Specific requirements for efficient and reproducible ISH investigations are discussed. Cells expressing the PPA gene in the adrenal medulla and in the brain were detected by ISH. The results show that ISH is as sensitive as immunohistochemistry in detecting peptide-producing cells in the adrenal and that it allows detection of PPA cell bodies in brain in conditions in which they are inconstantly detected by immunohistochemistry. Unilateral destruction of substantia nigra provokes a dramatic decrease in the number of neurons expressing the PPA gene in the contralateral striatum. Cells expressing the POMC gene were detected in the pituitary of various species including man and in the rat arcuate nucleus. Neurons containing vasopressin mRNA were visualized in the supraoptic paraventricular and suprachiasmatic nucleus of the adult rat by using a synthetic oligonucleotide probe. Transferrin gene expression was shown in the central nervous system of the rat brain in two cell populations, the oligodendrocytes and the epithelial cells of the choroid plexus, by demonstration of simultaneous presence in them of transferrin immunoreactivity together with transferrin mRNA. These results show that the ISH procedure is a technique that can be routinely used to investigate gene transcription anatomically in complex heterocellular tissues such as the endocrine glands and the nervous system.

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Section: Study Of Gene Expressionsupporting

confidence: 90%

In situ hybridization histochemistry for the analysis of gene expression in the endocrine and central nervous system tissues: A 3‐year experience

Bloch

Popovici

Guellec

et al. 1986

J of Neuroscience Research

130

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“…Oligonucleotide probes labeled on the 5' end with 32P have proved to be useful in in situ hybridization studies of vasopressin mRNA (34,(42)(43)(44)(45)(46)(47) and somatostatin mRNA (48), as well as POMC mRNA as reported here. This standard procedure for end-labeling oligonucleotides may not be adequate for routinely achieving fine cellular localization, especially when the mRNA abundance is low.…”

Section: Resultsmentioning

confidence: 99%

Detection of proopiomelanocortin mRNA by in situ hybridization with an oligonucleotide probe.

Lewis

Sherman

Burke

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

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Synthetic oligonucleotide probes can be easily obtained and used, in contrast to cDNA cloning to develop probes, and thus the present study was carried out to determine whether such probes could also be useful for in situ hybridization. A 24-base synthetic oligonucleotide complementary to part of the a-melanocyte-stimulating hormone (a-MSH) coding region of proopiomelanocortin (POMC) mRNA was 5'-end-labeled by using [y-32P]ATP with T4 polynucleotide kinase or was 3' tailed by using [a-32P] (200-250 g; Charles River Breeding Laboratories) were anesthetized with sodium pentobarbital (150 mg/kg, i.p.) and perfused intracardially with 50 ml of ice-cold saline followed by cold 0.1 M phosphate-buffered 4% formaldehyde for 30 min (21). One group of animals (n = 6) had been treated with haloperidol (2 mg/kg per day, i.p.) or saline for 4 days prior to perfusion. Pituitaries were removed, postfixed in ice-cold buffered formaldehyde for 1 hr, and incubated overnight in 15% sucrose in 0.02 M phosphate-buffered saline (21) at 4°C. The tissues were then frozen in liquid N2, and 10-,um-thick sections were cut in a cryostat at -20°C, thaw-mounted onto gelatin-coated slides, and stored at -80°C.Probes

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“…Among papers published previously (7,12,15,25,28,34,36,44,45), various conditions concerning hybridization and washing were employed in in situ hybridization with synthetic oligo-DNA probes. These situations were made more complicated by the fact that the theoretical formula does not always work in this area (2).…”

Section: Discussionmentioning

confidence: 99%

“…Most of the studies with oligo-DNA probes used 32P-labelling (12,28,34,36,44) or 1251-labelling (7) for signal detection though the resolution was not always sufficient for the analysis of individual cells. Recently, in order to obtain a better resolution and to minimize the cumbersome procedures associated with the handling of radioactive compounds, non-radioactive labels have been developed (13,19,26,35,42), and one of them has been applied to the in situ hybridization with oligo-DNA probe (15,25).…”

mentioning

confidence: 99%

In situ localization of c-myc mRNA in HL-60 cells using non-radioactive synthetic oligodeoxynucleotide probes.

Koji

Sugawara

Kimura

et al. 1989

Acta Histochem. Cytochem

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In situ hybridization of the vasopressin mRNA in the rat hypothalamus by use of a synthetic oligonucleotide probe

Cited by 42 publications

References 6 publications

In situ hybridization histochemistry for the analysis of gene expression in the endocrine and central nervous system tissues: A 3‐year experience

In situ hybridization histochemistry for the analysis of gene expression in the endocrine and central nervous system tissues: A 3‐year experience

Detection of proopiomelanocortin mRNA by in situ hybridization with an oligonucleotide probe.

In situ localization of c-myc mRNA in HL-60 cells using non-radioactive synthetic oligodeoxynucleotide probes.

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