Probing neurochemical structure and function of retinal ON bipolar cells with a transgenic mouse.
Retinal ON bipolar cells make up about 70% of all bipolar cells. Glutamate hyperpolarizes these cells by binding to the metabotropic glutamate receptor mGluR6, activating the G-protein G(o1), and closing an unidentified cation channel. To facilitate investigation of ON bipolar cells, we here report on the production of a transgenic mouse (Grm6-GFP) in which enhanced green fluorescent protein (EGFP), under control of mGluR6 promoter, was expressed in all and only ON bipolar cells. We used the mouse to determine density of ON bipolar cells, which in central retina was 29,600 cells/mm(2). We further sorted the fluorescent cells and created a pure ON bipolar cDNA library that was negative for photoreceptor unique genes. With this library, we determined expression of 27 genes of interest. We obtained positive transcripts for G(o) interactors: regulators of G-protein signaling (RGS), Ret-RGS1 (a variant of RGS20), RGS16, RGS7, purkinje cell protein 2 (PCP2, also called L7 or GPSM4), synembryn (RIC-8), LGN (GPSM2), RAP1GAP, and Gbeta5; cGMP modulators: guanylyl cyclase (GC) 1alpha1, GC1beta1, phosphodiesterase (PDE) 1C, and PDE9A; and channels: inwardly rectifying potassium channel Kir2.4, transient receptor potential TRPC2, and sperm-specific cation channels CatSper 2-4. The following transcripts were not found in our library: AGS3 (GPSM1), RGS10, RGS19 (GAIP), calbindin, GC1alpha2, GC1beta2, PDE5, PDE2A, amiloride-sensitive sodium channel ACCN4, and CatSper1. We then localized Kir2.4 to several cell types and showed that, in ON bipolar cells, the channel concentrates in their dendritic tips. The channels and modulators found in ON bipolar cells likely shape their light response. Additional uses of the Grm6-GFP mouse are also discussed.
Dhingra
Anuradha
A
Department of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6058, USA.
Sulaiman
Pyroja
P
Xu
Ying
Y
Fina
Marie E
ME
Veh
Rüdiger W
RW
Vardi
Noga
N
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D020869:Gene Expression Profiling; D015723:Gene Library; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D018094:Receptors, Metabotropic Glutamate / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D051245:Retinal Bipolar Cells / Q000166:cytology / Q000378:metabolism*; D015293:Transducin / Q000235:genetics / Q000378:metabolism
C489465:Kir2.4 channel; D024661:Potassium Channels, Inwardly Rectifying; D021122:Protein Subunits; D018094:Receptors, Metabotropic Glutamate; D011993:Recombinant Fusion Proteins; C113897:metabotropic glutamate receptor 6; C075192:gustducin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D015293:Transducin