Antisense probes against mediatophore block transmitter release in oocytes primed with neuronal mRNAs.
Antisense oligodesoxynucleotides were used to determine whether the mediatophore proteolipid is necessary for the Ca(2+)-dependent release of the neurotransmitter acetylcholine. Xenopus laevis oocytes were injected with poly(A)+ mRNAs extracted from the electric lobes of Torpedo marmorata. The electric lobes contain an homogeneous population of cholinergic neurons homologous to motoneurons. Addition of antisense probes hybridizing to the mediatophore 15 kDa subunit inhibited the expression of both the mediatophore proteolipid in oocyte membranes and the Ca(2+)-dependent acetylcholine release. Expression of other neuronal functions such as synthesis of [14C]acetylcholine from [14C]acetate was not inhibited. Another antisense probe specific for the sequence of a related proteolipid cDNA (the 15 kDa subunit of the chromaffin granule protonophore) was used as a control. It did not hybridize with the Torpedo mediatophore mRNA and, injected in addition to electric lobe mRNAs, it did not inhibit either mediatophore expression or acetylcholine release. We showed in addition that the mRNA primed oocytes did not contain a vesicular pool of acetylcholine. It was concluded (i) that the mediatophore proteolipid is essential for Ca(2+)-dependent acetylcholine release and (ii) that the cytosolic pool of neurotransmitter seems to be preferentially used in this system.
Cavalli
A
A
Département de Pharmacologie, CMU, Genève, Switzerland.
Dunant
Y
Y
Leroy
C
C
Meunier
F M
FM
Morel
N
N
Israël
M
M
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D016375:Antisense Elements (Genetics) / Q000494:pharmacology*; D001483:Base Sequence; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D002837:Chromaffin Granules / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry; D004557:Electric Organ / Q000378:metabolism*; D005260:Female; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009046:Motor Neurons / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D009474:Neurons / Q000378:metabolism*; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D011061:Poly A / Q000378:metabolism; D011510:Proteolipids / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D012333:RNA, Messenger / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D014101:Torpedo; D014982:Xenopus laevis
D016375:Antisense Elements (Genetics); D018076:DNA, Complementary; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D011510:Proteolipids; D012333:RNA, Messenger; C050299:mediatophore; D011061:Poly A; D000109:Acetylcholine; D002118:Calcium