Characterization of recombinant and native Ih-channels from Apis mellifera.
Recently, a novel class of genes coding for Ih-channels has been identified in several vertebrates and invertebrates. We isolated a cDNA (AMIH) encoding a putative member of these ion channels from Apis mellifera heads by means of polymerase chain reaction and homology screening. High similarity (88% identical amino acids) to the putative Drosophila melanogaster Ih-channel suggests that the Apis cDNA codes for a hyperpolarization-activated and cyclic nucleotide-gated channel. Functional expression of recombinant AMIH in HEK293 cells gave unitary currents that were preferentially selective for potassium over sodium ions and were activated by hyperpolarizing voltage steps. Cyclic nucleotides shifted the voltage activation curve to more positive membrane potentials. The current kinetics, activation by hyperpolarizing voltage steps and modulatory influence of cyclic nucleotides properties closely resemble those of mammalian Ih-channels. RT-PCR analysis showed pronounced mRNA expression in the antennae, head and body of Apis mellifera. Investigation of hyperpolarization-activated currents in olfactory receptor neurons (ORNs) in a primary cell culture of Apis mellifera antennal cells revealed activation properties similar to the heterologously expressed Ih-channel. By in-situ hybridization and immunohistochemistry, expression of AMIH was seen in olfactory receptor neurons of the bee antennae. We conclude that AMIH is the ion channel responsible for the hyperpolarization-activated currents in olfactory receptor neurons of bee.
Gisselmann
Günter
G
Fakultät für Biologie, Lehrstuhl für Zellphysiologie ND4, Ruhr-Universität-Bochum, Universitätsstr 150, 44780 Bochum, Germany. guenter.gisselmann@ruhr-uni-bochum.de
Warnstedt
Maike
M
Gamerschlag
Birgit
B
Bormann
Anja
A
Marx
Thomas
T
Neuhaus
Eva M
EM
Stoertkuhl
Klemens
K
Wetzel
Christian H
CH
Hatt
Hanns
H
D000595:Amino Acid Sequence; D000818:Animals; D001516:Bees / Q000166:cytology / Q000235:genetics* / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D002586:Cesium / Q000494:pharmacology; D003001:Cloning, Molecular; D054815:Cyclic Nucleotide-Gated Cation Channels; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D004594:Electrophysiology / Q000379:methods; D015870:Gene Expression; D017344:Genes, Insect; D006801:Humans; D007473:Ion Channels / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D018034:Olfactory Receptor Neurons / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D054815:Cyclic Nucleotide-Gated Cation Channels; D018076:DNA, Complementary; D007473:Ion Channels; D011994:Recombinant Proteins; D002586:Cesium