A floxed transcription-disrupter (TD) ablates expression of the mouse Hcrtr1 gene in this OX1R TD conditional mutant strain. Cre-mediated excision of the TD restores expression under the direction of the native promoter.
Thomas Scammell, Beth Israel Deaconess Medical Center
The Hcrtr1 (hypocretin (orexin) receptor 1; also called OX1R) gene plays a role in modulating reward and depression-like behavior. Orexin signaling has been implicated in processes as diverse as sleep-wake transitioning, the control of food intake, and autonomic output.
In this conditional mutant OX1R TD strain, a loxP-flanked transcription-disrupter (TD) cassette was introduced to intron 1 of the mouse Hcrtr1 gene to prevent expression of functional protein. Cre-mediated excision of the TD restores expression under the direction of the native promoter.
The impact of this mutation was first tested electrophysiologically using in vitro patch-clamp recordings of noradrenergic neurons of the locus coeruleus (LC), a type of neuron that expresses only OX1R but not OX2R. Orexin-A ligand depolarizes LC neurons from wildtype mice, but has no effect on LC neurons from homozygous OX1R TD mice. The deficit in LC neurons is fully restored after crossing OX1R TD mice with Zp3-Cre mice, which express Cre recombinase in the female germline.
Behaviorally, OX1R TD mice have a significant reduction in behavioral despair in the forced swim test and tail suspension test, but anxiety-like behavior is normal. These mice have normal wake, non-REM, and REM sleep behavior. Cataplexy (episodes of sudden muscle atonia during active wakefulness) does not occur in these mice. OX1/2R TD progeny from crosses with OX2R TD mice (see Stock No. 027707) have severely fragmented sleep/wake behavior, similar to that seen in prepro-orexin null mice.
OX1R TD mice are useful as OX1R null mice in their native state. More importantly, they can be used to study the effects of focal or global restoration of OX1R signaling, using methods that produce focal or germline expression of Cre recombinase, respectively. Focal OX1R restoration can be used to test whether OX1R signaling in a specific brain region is sufficient to rescue specific behaviors.
A floxed transcription disrupter cassette was inserted into intron 1 (formerly identified as intron 3) via homologous recombination in 129-derived embryonic stem (ES) cells. The TD cassette consists of a mouse En2 splice acceptor site followed by an SV40 enhancer, neomycin resistance gene, and two HSV-TK poly(A) signals (a synthetic poly(A) signal/transcriptional pause signal, and another synthetic poly(A) signal followed by a Myc-associated zinc finger protein-binding site). Resultant founder mice were backcrossed to C57BL/6J mice for more than 15 generations by the donating laboratory.
|Allele Name||targeted mutation 1, Michael M Scott|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Gene Symbol and Name||Hcrtr1, hypocretin (orexin) receptor 1|
|Strain of Origin||129|
|Molecular Note||A floxed transcription disrupter cassette was inserted into intron 3 via homologous recombination. In situ hybridization analysis confirmed the absence of expression in the central nervous system of homozygous mice.|
Homozygotes and heterozygotes are viable and fertile.
When using the Hcrtr1 null mouse strain in a publication, please cite the originating article(s) and include JAX stock #027706 in your Materials and Methods section.