Korean J Anat.  1998 Dec;31(6):807-815.

Study on the Expression of Vasoactive intestinal polypeptide, Vasopressin and Oxytocin mRNAs in the Rat Brain using Double in situ Hybridization Technique

• Park KH
• Park HS
• Yoo YB
• Lee WJ
• Hwang DH
• Lee BL
• Cha CI
• Cho SS
• Baik SH

Abstract

The technique of in situ hybridization using synthetic oligonucleotides labelled by non-radioactive method was developed to localize vasoactive intestinal polypeptide, arginine-vasopressin and oxytocin mRNAs in the rat brain. Also double in situ hybridization technique where combination of non-radioactive and radioactive probes were applied was developed to localize 2 neuropeptide mRNAs in single tissue section. The results were as follows; In non-radioactive in situ hybridization methods using digoxigenin-labelled oligonucleotide probe, alkaline-phosphates method using NBT and BCIP as substrates gave the best result that specific hybridization signals were observed. In radioactive in situ hybridization methods using 35S-labelled oligonucleotide probe, specific hybridization signals were observed in both nuclear track emulsion and X-ray film autoradiography. In double in situ hybridization methods using combination of 35S-labelled and digoxigenin-labelled oligonucleotide probes, specific hybridization signals were observed in the group where K5 emulsion was applied as nuclear track emulsion. The technique of in situ hybridization using digoxigenin-labelled oligonucleotide applied in this study will be useful as alternative for radioactive in situ hybridization technique. Moreover, combination of non-radioactive and radioactive labelled probes in double in situ hybridization technique will be a useful tool for the simultaneous localization of various mRNAs in single section for the study of various neurotransmitters, neuropeptides, receptors and signal transduction molecules.

Keyword

In situ hybridization; Double labelling; Digoxigenin; Radioisotope; Vasoactive intestinal polypeptide; Vasopressin; Oxytocin; Aging; Gender; Corpus callosum; Midbrain; Pons; Cerebellar vermis; MRI

MeSH Terms

Aging
Animals
Autoradiography
Brain*
Corpus Callosum
Digoxigenin
In Situ Hybridization*
Magnetic Resonance Imaging
Mesencephalon
Neuropeptides
Neurotransmitter Agents
Oligonucleotide Probes
Oligonucleotides
Oxytocin*
Pons
Rats*
RNA, Messenger*
Signal Transduction
Vasoactive Intestinal Peptide*
Vasopressins*
X-Ray Film
Digoxigenin
Neuropeptides
Neurotransmitter Agents
Oligonucleotide Probes
Oligonucleotides
Oxytocin
RNA, Messenger
Vasoactive Intestinal Peptide
Vasopressins