Detection of mRNA molecules coding for neuropeptide hormones of the pond snail Lymnaea stagnalis by radioactive and non-radioactive in situ hybridization: a model study for mRNA detection

Journal Article · Sun Jan 01 00:00:00 EST 1989 · J. Histochem. Cytochem.; (United States)

DOI:https://doi.org/10.1177/37.1.2642295· OSTI ID:6611747

Dirks, R W; Raap, A K; Van Minnen, J; Vreugdenhil, E; Smit, A B; Van der Ploeg, M

To develop and optimize non-radioactive in situ hybridization techniques for mRNA detection, we used the neuropeptidergic system of the pond snail Lymnaea stagnalis as a biological model system. First, we investigated the in situ hybridization procedure using radioactive-labeled cDNA and synthetic oligonucleotide probes specific for egg-laying hormone (ELH) mRNA and molluscan insulin-like peptide (MIP) mRNA. The results show an intense grain deposit above the caudodorsal cells and light-green cells expressing, respectively, ELH mRNA and MIP mRNA. Good results with relation to signal strength and tissue morphology were obtained with freeze-dry paraformaldehyde vapor fixation. The necessity to perform tissue pre-treatment appeared to be dependent on the cell type of interest. The optimized in situ hybridization protocol proved to be applicable using probes that are either sulfonated/transaminated or labeled with acetylaminofluorene (AAF). In situ hybridization of such haptenized probes led to intense and specific staining of the cytoplasm of the caudodorsal cells. Egg-laying hormone mRNA appeared not to be homogeneously distributed in the cytoplasm but showed a patch-like pattern. Nuclear and axoplasmic staining for mRNA was also observed.

• Research Organization: Univ. of Leiden (Netherlands)
• OSTI ID: 6611747
• Journal Information: J. Histochem. Cytochem.; (United States), Vol. 37:1
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
MESSENGER-RNA
HYBRIDIZATION
BIOLOGICAL MODELS
CYTOPLASM
DNA
INSULIN
OLIGONUCLEOTIDES
PEPTIDES
RADIOISOTOPES
SNAILS
STAINS
TRACER TECHNIQUES
ANIMALS
AQUATIC ORGANISMS
CELL CONSTITUENTS
HORMONES
INVERTEBRATES
ISOTOPE APPLICATIONS
ISOTOPES
MOLLUSCS
NUCLEIC ACIDS
ORGANIC COMPOUNDS
PEPTIDE HORMONES
PROTEINS
RNA
550201* - Biochemistry- Tracer Techniques