Mahoney, A. | bioRxiv Science

bioRxiv2023

An osmolarity dependent mechanism partially ameliorates retinal cysts and rescues cone function in a mouse model of X-linked Retinoschisis.

IntroductionX-linked retinoschisis (XLRS) is a vitreoretinal dystrophy caused by RS1 gene mutations which disrupt retinoschisin protein function. A vital protein for maintaining retinal architecture, the absence of functional retinoschisin leads to the development of intraretinal cysts. The preliminary goal of this study was to investigate a low dose gene therapy in Rs1 knockout (Rs1-KO) mice; however, our experiments revealed an unexpected therapeutic effect of a hypertonic buffer, which led to further exploration of this effect. Methods10 Rs1-KO mice were subretinally injected with an AAV2/4 vector containing the RS1 gene driven by an Ef1 promoter. 16 Rs1-KO mice were subretinally injected with a hypertonic buffer (180 mM NaCl 0.001% F68/PBS (pH 7.4)) or an isotonic buffer (155.2 mM NaCl 0.001% F68/PBS, pH 7.0) as a sham control. Endpoints included electroretinogram (ERG), optical coherence tomography (OCT), and a visually guided swim assay (VGSA). An immunohistochemistry assay was used to quantify cone density in buffer injected and treatment-naive eyes. ResultsUnexpectedly, hypertonic buffer-injected eyes had significantly reduced cyst severity at 1 month post-injection (MPI) (p=<0.0001), significantly higher amplitudes in cone-dominant ERGs persisting to 5 months post-injection (5 Hz flicker; p=0.0018; 3.0 Flash; p=0.0060) and demonstrated improved navigational vision in the light compared to untreated Rs1-KO eyes (p<0.0001). To investigate the role of tonicity on this effect, an isotonic buffer-injected cohort was created (155.2 mM NaCl 0.001% F68/PBS, pH 7.0) (n=6). Surprisingly, hypertonic buffer-injected eyes exhibited a greater reduction in cyst severity and demonstrated improved cone-dominant ERG metrics over isotonic buffer-injected eyes. Using an immunohistochemistry assay, we demonstrated greater cone density in hypertonic buffer-injected eyes than untreated controls (p=0.0147), suggesting a possible cone preservation mechanism. Moreover, our findings reveal a negative correlation between the peak severity of cysts and long-term cone-dominant ERG metrics, implying that effectively managing cysts could yield enduring benefits for cone function. Discussion/ConclusionThis study presents evidence that cyst resolution can be triggered through an osmosis-dependent pathway, and cyst resolution can have long term effects on cone signaling and survival, offering potential insights for the development of novel treatments for patients with XLRS.

genetics↗

bioRxiv2021

Delineation of dynorphin and kappa opioid receptor circuit elements in alcohol consumption

Alcohol use disorder is complex and multi-faceted, involving the coordination of multiple signaling systems across numerous brain regions. Previous work has indicated that both the insular cortex and dynorphin (DYN)/Kappa opioid receptor (KOR) systems contribute to excessive alcohol use. More recently, we identified a microcircuit in the medial aspect of the insular cortex that signals through DYN/KOR. Here, we explored the role of insula DYN/KOR circuit components on alcohol intake in a long-term intermittent access (IA) procedure. Using a combination of conditional knockout strategies and site-directed pharmacology, we discovered distinct and sex-specific roles for insula DYN and KOR in alcohol drinking and related behavior. Our findings show that insula DYN deletion blocked escalated consumption and decreased overall intake of and preference for alcohol in male and female mice. This effect was specific to alcohol in male mice, as DYN deletion did not impact sucrose intake. Further, insula KOR antagonism reduced alcohol intake and preference during the early phase of IA in male mice only. Alcohol consumption was not affected by insula KOR knockout in either sex. In addition, we found that long-term IA decreased the intrinsic excitability of DYN and deep layer pyramidal neurons (DLPN) in the insula of male mice. Excitatory synaptic transmission was also impacted by IA, as it drove an increase in excitatory synaptic drive in both DYN neurons and DLPN. Combined, our findings suggest there is a dynamic interplay between excessive alcohol consumption and insula DYN/KOR microcircuitry. Significance StatementThe insular cortex is a complex region that serves as an integratory hub for sensory inputs. In our previous work, we identified a microcircuit in the insula that signals through the kappa opioid receptor (KOR) and its endogenous ligand dynorphin (DYN). Both the insula and DYN/KOR systems have been implicated in excessive alcohol use and alcohol use disorder (AUD). Here, we utilize converging approaches to determine how insula DYN/KOR microcircuit components contribute to escalated alcohol consumption. Our findings show that insula DYN/KOR systems regulate distinct phases of alcohol consumption in a sex-specific manner, which may contribute to the progression to AUD.

neuroscience↗