Biology subjects

Wellington, A. J.

Publications and source records attributed to Wellington, A. J..

3 recordsLinked to original sources

bioRxiv2025

Early Diabetic Ca2+ Handling Impairments in the Rod Bipolar Pathway

BackgroundPrevious work showed that electrically-evoked inhibition to Rod Bipolar Cells (RBC) is reduced in a mouse model of early diabetes. It is hypothesized that this is due to impaired Ca2+ handling in the presynaptic amacrine cell, either through increased Ca2+ buffering or decreased influx. To test this hypothesis and develop a mechanism for this effect, a model where direct optogenetic activation of inhibitory amacrine cells that expressed the light-activated channel ChR2 was used to isolate amacrine cell inputs to RBCs. Application of selective Ca2+ channel blockers could then assess potential locations of amacrine Ca2+ disruption. Using whole cell patch clamp electrophysiology, recordings were made from a 6 week diabetic population (DM) and vehicle injected non-DM animals. ResultsRobust GABAC receptor inhibitory currents were recorded from RBCs after ChR2 stimulus that were significantly diminished by the application of nifedipine to block L-type Ca2+ channels in both DM and non-DM conditions. There were significant differences in the peak amplitude of these responses between DM and non-DM groups (p = 0.0146). However, in the non-DM group the decay tau of the response to the 50ms stimulus was significantly diminished by nifedipine ({tau} p =0.0498, n = 5), but this was not seen in the DM group ({tau} p = 0.9498, n=7). A 1s nifedipine-reduced response saw its decay tau increase in the DM group but not the non-DM. Ca2+ - induced Ca2+ release (CICR) blockade with ryanodine decreased responsivity equally between groups in the 1s stimulus but showed no significant kinetic changes. CICR blockade for a 50ms stimulus response showed significant kinetic changes in diabetes but otherwise reduced the response equally between DM and non-DM. Blockade of the mitochondrial Ca2+ uniporter (MCU) had little effect on the optogenetic response. ConclusionThis study presents evidence that diabetes alters amacrine cell output to the RBC unmasked through blockade of the L-type calcium channel, and the Endoplasmic Reticulum (ER). An apparent explanation for our results is that DM calcium buffering is dysregulated, leading to prolonged responses. The underlying mechanism for this alteration is complex and not yet clearly elucidated.

neuroscience↗

bioRxiv2025

Dopamine D1 Receptors Modulate glycinergic Inhibitory Inputs to OFF cone bipolar cells elicited by Optogenetic Activation of Amacrine Cells

PurposeLight evoked inhibition of OFF cone bipolar cells (OFF BCs) is modulated both by background light levels and the action of dopamine through the dopamine D1 receptor (R). Since D1Rs are localized throughout the mouse retina, it is not known where in the light signaling pathway dopamine is modulating signals to OFF BCs. Here we tested a technique that allowed for the isolation of the amacrine cell (AC) to OFF BC circuit to determine if there are local D1R-induced changes in inhibition from presynaptic ACs onto OFF BCs. MethodsWe utilized the B6.Cg-Tg(Slc32al-COP4*H134R/EYFP) mouse line that expresses ChR2 in all the inhibitory cells in the retina. ACs expressing ChR2 were directly activated by light, while blocking photoreceptor mediated inputs. Inhibitory synaptic currents or ChR2-evoked excitatory currents were recorded using whole-cell patch clamp electrophysiology. ResultsOptogenetically activated inhibition from ACs elicited inhibitory currents that had slow kinetics similar to light-evoked inhibition, suggesting that they are using native synaptic mechanisms. We recorded optogenetically activated inputs to OFF BCs from pharmacologically isolated GABAergic and glycinergic input while photoreceptor inputs were blocked. D1R activation reduced glycinergic inputs to OFF BCs while leaving GABAergic inputs intact. ConclusionsD1R modulation of isolated optogenetic activation of AC-OFF BC synapses showed similar changes to previous experiments with light-evoked inhibition to OFF BCs. Together, this suggests optogenetic activation of ACs can be used to understand how dopamine differentially shapes inhibitory changes in the inner retina.

neuroscience↗

bioRxiv2020

Impaired light adaptation of ON-sustained ganglion cells in early diabetes is attributable to diminished dopamine D4 receptor sensitivity

Purpose: It has been known for some time that normal retinal signaling is disrupted early on in diabetes, before the onset of the vascular pathologies associated with diabetic retinopathy. There is growing evidence that levels of retinal dopamine, a neuromodulator that mediates light adaptation, may also be reduced in early diabetes. Previously, we have shown that after six weeks of diabetes in a mouse model, light adaptation is impaired at the level of ON-sustained (ON-s) ganglion cells. The purpose of this study was to determine whether changes in dopamine receptor sensitivity contribute to this dysfunction. Here we used single cell retinal patch-clamp recordings from the mouse retina to determine how activating dopamine type D4 receptors (D4Rs) changes the light-evoked and spontaneous excitatory inputs to ON-s ganglion cells, in both control and diabetic animals. We also used in-situ fluorescent hybridization to assess whether D4R expression was impacted by diabetes. We found that D4R activation had a smaller impact on light-evoked excitatory inputs to ON-s ganglion cells in diabetic retinas compared to controls. This impaired D4R signaling is not attributable to a decline in D4R expression, as we found increased D4R mRNA density in the outer plexiform layer in diabetic retinas. This suggests that the cellular machinery of dopaminergic signaling is itself disrupted in early diabetes and may be amenable to chronic dopamine supplementation therapy.

neuroscience↗