Genomic and functional conservation of sedative-hypnotic targets in the zebrafish.
OBJECTIVE
The zebrafish is an ideally suited vertebrate animal model for large-scale genetic screens and is emerging as a model organism in pharmacological and behavioral research. We investigated the effects of sedative hypnotics commonly used in humans on zebrafish locomotor activity and identified the corresponding genomic and receptor binding targets.


METHODS
We studied radioreceptor binding and behavioral responses to compounds with known sedative hypnotic properties representing multiple pharmacological classes. These included GABAergic hypnotics such as benzodiazepines, barbiturates, and baclofen; alpha-2 adrenergic agonists; and histaminergic H1 antagonists. An automated system was used to quantify behavioral effects. Zebrafish homologs of histamine receptor H1, gamma-amino-n-butyric acid type A (alpha-subunit), and gamma-amino-n-butyric acid type B (1 and 2) receptor genes were identified through translating queries of the zebrafish Zv4 database with human receptor protein sequences. A pilot screen of 154 N-ethyl-N-nitroso-urea-mutagenized F2 families was conducted with pentobarbital, flurazepam and mepyramine.


RESULTS
Radioreceptor binding studies revealed high affinity binding sites for known gamma-amino-n-butyric acid type A, gamma-amino-n-butyric acid type B, and histaminergic ligands. Drug immersion of 5-7-day-old larvae reduced mobility and, in some cases, produced a complete state of unresponsive immobility similar to anesthesia. These effects were dose-dependent and rapidly reversible in water. As established in mammals, (R)-baclofen was more active behaviorally and had higher affinity in binding studies when compared with (S)-baclofen. In this model, (S)-baclofen only partially reduced activity at high dose and blocked (R)-baclofen behavioral hypnotic effects. Genomic sequences with high similarity to the corresponding pharmacological targets were identified, but no mutants were found in the pilot screen.


CONCLUSIONS
These results demonstrate conservation of gene, protein and function for many established sedative hypnotic pathways. The results indicate feasibility of conducting large-scale pharmacogenomic screens to isolate novel proteins modulating susceptibility to hypnotic compounds in a vertebrate system.
Renier
Corinne
C
Department of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, Palo Alto, California, USA.
Faraco
Juliette H
JH
Bourgin
Patrice
P
Motley
Timothy
T
Bonaventure
Pascal
P
Rosa
Frédéric
F
Mignot
Emmanuel
E
D000595:Amino Acid Sequence; D000818:Animals; D001418:Baclofen / Q000737:chemistry / Q000494:pharmacology; D001522:Behavior, Animal / Q000187:drug effects; D001665:Binding Sites / Q000235:genetics; D017124:Conserved Sequence; D006801:Humans; D006993:Hypnotics and Sedatives / Q000737:chemistry / Q000494:pharmacology*; D007814:Larva / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D008969:Molecular Sequence Data; D009043:Motor Activity / Q000187:drug effects / Q000235:genetics; D010597:Pharmacogenetics; D010802:Phylogeny; D011869:Radioligand Assay; D011963:Receptors, GABA-A / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D018080:Receptors, GABA-B / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D011969:Receptors, Histamine H1 / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015027:Zebrafish / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*
D006993:Hypnotics and Sedatives; D011963:Receptors, GABA-A; D018080:Receptors, GABA-B; D011969:Receptors, Histamine H1; D001418:Baclofen