Abstract
Null mice for the dopamine D2 receptor (D2R) have been instrumental in understanding the function of this protein. For our research, we obtained the functional D2R knockout mouse strain described initially in 1997. Surprisingly, our biochemical characterization showed that this mouse strain is not a true knockout. We determined by sequence analysis of the rapid 3′ amplification of cDNA ends that functional D2R knockout mice express transcripts that lack only the eighth exon. Furthermore, immunofluorescence assays showed a D2R-like protein in the brain of functional D2R knockout mice. We verified by immunofluorescence that the recombinant truncated D2R is expressed in HEK293T cells, showing intracellular localization, colocalizing in the Golgi apparatus and the endoplasmic reticulum, but with less presence in the Golgi apparatus compared to the native D2R. As previously reported, functional D2R knockout mice are hypoactive and insensitive to the D2R agonist quinpirole. Concordantly, microdialysis studies confirmed that functional D2R knockout mice have lower extracellular dopamine levels in the striatum than the native mice. In conclusion, functional D2R knockout mice express transcripts that lead to a truncated D2R protein lacking from the sixth transmembrane domain to the C-terminus. We share these findings to avoid future confusion and the community considers this mouse strain in D2R traffic and protein–protein interaction studies.
Highlights
Dopamine is a neurotransmitter that participates in the control of voluntary movements and motivated behaviors among other relevant functions
The D2 receptors (D2R) functional knockout mouse strain was generated by Kelly et al (1997) by inserting a neomycin cassette into the genomic site corresponding to exon 7 and most of exon 8 according to the mouse genome description available at that time [11]
The recharacterization of the functional D2R knockout mouse strain performed in our work shows that these mice are better described as D2R knock-in mice, in which the Drd2 gene has a deletion of exon 8
Summary
Dopamine is a neurotransmitter that participates in the control of voluntary movements and motivated behaviors among other relevant functions. Two types of seventransmembrane segment receptors mediate the action of dopamine, type 1 receptors coupled to the excitatory G protein (Gs) which include D1 and D5 receptors, and type 2 receptors coupled to the inhibitory G protein (Gi/o) which include D2, D3, and D4 receptors. Cumulative evidence indicates that the D2L variant mediates mainly dopamine postsynaptic actions, whereas the D2S variant mediates presynaptic control of dopamine release and dopamine neuron firing [3]. D2R are especially abundant in the striatopallidal efferent GABA medium spiny neurons (MSN) that are involved in the control of voluntary movements [5,6]. The activation of postsynaptic D2R in these GABA efferent MSN induces locomotor activity by decreasing
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