Journal: bioRxiv

Article Title: Dyrk1a gene dosage controls bipolar cell development and retinal connectivity

doi: 10.64898/2026.03.15.710015

Figure Lengend Snippet: (A) Schematic representation of electroretinogram (ERG) components. An initial negative deflection (a-wave) is observed upon photoreceptor stimulation (arrow) that is followed by a subsequent positive deflection (b-wave) that corresponds to bipolar cell activity. ERG waveform quantification involves measuring the amplitude (μV) and latency (ms) of the a- and b-waves to increasing stimulus intensities (cd.s/m 2 ) and thus provides an evaluation of retinal function. (B) Representative dark-adapted scotopic ERG waveforms recorded from Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animals (6 – 8 weeks) in response to a single flash stimulus (log 1.48 cd.s/m 2 ). Comparable a-wave responses are observed for both groups while b-wave amplitudes are reduced in heterozygous animals. Thick lines represent genotype-mean responses while thin lines represent individual mice. (C – D) Quantifcation of photoreceptor amplitude (C) and latency (D) responses in Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animals (6 – 8 weeks) in repsonse to a series of increasing scotopic flash stimuli (log cd.s/m 2 ). No significant differences are observed between the groups. (E – F) Quantification of bipolar cell amplitude (E) and latency (F) responses in Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animal (6 – 8 weeks) towards increasing scotopic flash stimuli (log cd.s/m 2 ). Heterozygous animals have a significant reduction in bipolar cell amplitude response at higher stimulus intensities (E) albeit with similar latency (F). (G) Representative light-adapted photopic ERG waveforms recorded from Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animals (6 – 8 weeks) in response to a single UV flash stimulus (log 1.00 cd.s/m 2 ). Comparable a-wave responses are observed for both groups while b-wave amplitudes are reduced in heterozygous animals. Thick lines represent genotype-mean responses while thin lines refer to individual mice. (H) Quantification of bipolar cell amplitude response in Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animal (6 – 8 weeks) in response to an increasing photopic UV stimulus intensity series (log cd.s/m 2 ). Heterozygous animals have a significant reduction in bipolar cell amplitude response at higher stimulus intensities. (I) Representative light-adapted photopic ERG waveforms recorded from Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animals (6 – 8 weeks) in response to a single green flash stimulus (log 1.48 cd.s/m 2 ). Comparable a-wave responses are observed for both groups while b-wave amplitudes are reduced in heterozygous animals. Thick lines represent genotype-mean responses while thin lines refer to individual mice. (J) Quantification of bipolar cell amplitude response in Lhx2-Cre ( n = 3) and Lhx2-Cre:Dyrk1a +/f ( n = 3) animal (6 – 8 weeks) in response to a photopic green stimulus intensity series (log cd.s/m 2 ). Heterozygous animals have a significant reduction in bipolar cell amplitude response at higher stimulus intensities. (K – R) Representative immunohistochemical staining of coronal eye sections taken from Lhx2-Cre and Lhx2-Cre:Dyrk1a +/f adult mice (6 – 8 weeks) across four designated dorsoventral quadrants (D1, D2, V3 and V4) spanning the entire retina relative to the optic nerve head. OPL ribbon synapse assemblies appear comparable in both groups (K – R, arrowheads) and were composed of dense photoreceptor terminals (vGlut1 + ) that overlapped with well organised bipolar cell dendritic processes (PKCα + ). Additionally, the linear alignment of all ribbon junctions (Pikachurin + ) indicated intact photoreceptor and bipolar cell synapses. All data represents the mean ± SEM. Statistical differences were calculated using a two-way ANOVA followed by a Sidak’s multiple comparison test. p-values are denoted as follows: **p≤ 0.01, ***p≤ 0.001 and ****p≤ 0.0001. Scale bar: (K – R) 5 µm. Abbreviations: D, dorsal; GCL, ganglion cell layer; INL, inner nuclear layer; IPL, inner plexiform layer; ONL, outer nuclear layer; OPL, outer plexiform layer; V, ventral.

Article Snippet: The following primary antibodies and dilutions were used in this study: Calbindin (1:300, Sigma-Aldrich, #C9848); Calretinin (1:1000, Swant, #CG1); cleaved Caspase3 (Asp175) (1:500, Cell Signalling, #9661S); chAT (1:100, Millipore, #AB144P); Chx10 (1:50, SCBT, #SC365519); Dyrk1a (1:100, Novus, #NBP1-84032); Isl1/2 (recognises both Isl1 and Isl2 proteins) (1:500; DSHB, #39.4D5); M-opsin (1:500, Proteintech, #30975-1-AP); Pax6 (1:100, DSHB, #Pax6); Pikachurin (1:200, Proteintech, #14578-1-AP); Rhodopsin (1:500, Millipore, #MABN15); S-opsin (1:500, Proteintech, #24660-1-AP); TH (1:300, Millipore, #AB152); vGlut1 (1:2000, Millipore, #AB5905).

Techniques: Activity Assay, Immunohistochemical staining, Staining, Comparison