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Background/aims: The authors have previously reported that carbonic anhydrase inhibitors such as acetazolamide and dorzolamide raise optic nerve oxygen tension (ONP[O.sub.2]) in pigs. The purpose of the present study was [...]

Presents various studies on genetics. Observance of interallic complementation at the mouse microphthalmia locus; Details on electroretinographic and histologic assessment of retinal function in an allelic series of microphthalmia mutant mice; Construction of gene transfer vector based on maedi-visna virus.

Wolfram syndrome (WS) is a rare condition caused by homozygous or compound heterozygous mutations in the WFS1 gene primarily. It is diagnosed on the basis of early-onset diabetes mellitus and optic nerve atrophy. Patients complain of trigeminal-like migraines and show deficits in vibration sensation, but the underlying cause is unknown. Using accurate cell models and two separate, accurate rodent models of WS that show excellent face and construct validity, here we have examined trigeminus, sensation and sensory neuronal function in WS. Analysis of ex vivo and in vivo MRI sequences revealed profound trigeminal atrophy in each rodent model, a novel finding in WS. Optic nerve atrophy is a diagnostic sign in WS, and trigeminal atrophy occurred at the time of earliest loss of optic nerve volume. We also observed deficits in mechanical sensation in our mouse WS model, and pathological analysis revealed extensive inflammation in trigeminal sensory nucleus, both of which are novel findings in WS. Sensory neurons (dorsal root ganglia) showed impaired calcium handling upon depolarisation and reduced mitochondrial membrane potential. Finally, lysosomes were smaller, soma lysosome content was decreased and importantly, lysosome acidity was impaired in sensory neurons, all of which are novel findings in WS. We validated these findings using two separate publicly available datasets, both from WS patient fibroblast-derived neural stem cells. We observed a highly significant functional enrichment of GO cellular component lysosome-related terms among the differentially expressed proteins and genes, with the majority of lysosome-related proteins being downregulated. These data reveal extensive impairments in the trigeminal pathway and nociceptive neurons in WS that may contribute to trigeminal and sensory symptoms observed in patients. Moreover, we note that mutations in WFS1 are relatively common and, given the importance of WFS1 for sensory function, our data may also shed light on sensory impairments in general., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background/objectives: The microphthalmia-associated transcription factor ( Mitf ) has been found to play an important role in eye development, structure, and function. The Mitf gene is responsible for controlling cellular processes in a range of cell types, contributing to multiple eye development processes. In this review, we survey what is now known about the impact of Mitf on eye structure and function in retinal disorders. Several mutations in the human and mouse Mitf gene are now known, and the effects of these on eye phenotype are addressed. We discuss the importance of Mitf in regulating ion transport across the retinal pigment epithelium (RPE) and the vasculature of the eye., Methods: The literature was searched using the PubMed, Scopus, and Google Scholar databases. Fundus and Optical Coherence Tomography (OCT) images from mice were obtained with a Micron IV rodent imaging system., Results: Defects in neural-crest-derived melanocytes resulting from any Mitf mutations lead to hypopigmentation in the eye, coat, and inner functioning of the animals. While many Mitf mutations target RPE cells in the eye, fewer impact osteoclasts at the same time. Some of the mutations in mice lead to microphthalmia, and ultimately vision loss, while other mice show a normal eye size; however, the latter, in some cases, show hypopigmentation in the fundus and the choroid is depigmented and thickened, and in rare cases Mitf mutations lead to progressive retinal degeneration., Conclusions: The Mitf gene has an impact on the structure and function of the retina and its vasculature, the RPE, and the choroid in the adult eye.

It has been known for some time that Carbonic Anhydrase (CA, EC 4.2.1.1) plays a complex role in vascular function, and in the regulation of vascular tone. Clinically employed CA inhibitors (CAIs) are used primarily to lower intraocular pressure in glaucoma, and also to affect retinal blood flow and oxygen saturation. CAIs have been shown to dilate vessels and increase blood flow in both the cerebral and ocular vasculature. Similar effects of CAIs on vascular function have been observed in the liver, brain and kidney, while vessels in abdominal muscle and the stomach are unaffected. Most of the studies on the vascular effects of CAIs have been focused on the cerebral and ocular vasculatures, and in particular the retinal vasculature, where vasodilation of its vessels, after intravenous infusion of sulfonamide-based CAIs can be easily observed and measured from the fundus of the eye. The mechanism by which CAIs exert their effects on the vasculature is still unclear, but the classic sulfonamide-based inhibitors have been found to directly dilate isolated vessel segments when applied to the extracellular fluid. Modification of the structure of CAI compounds affects their efficacy and potency as vasodilators. CAIs of the coumarin type, which generally are less effective in inhibiting the catalytically dominant isoform hCA II and unable to accept NO, have comparable vasodilatory effects as the primary sulfonamides on pre-contracted retinal arteriolar vessel segments, providing insights into which CA isoforms are involved. Alterations of the lipophilicity of CAI compounds affect their potency as vasodilators, and CAIs that are membrane impermeant do not act as vasodilators of isolated vessel segments. Experiments with CAIs, that shed light on the role of CA in the regulation of vascular tone of vessels, will be discussed in this review. The role of CA in vascular function will be discussed, with specific emphasis on findings with the effects of CA inhibitors (CAI)., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer SA declared a past co-authorship with the author CS to the handling editor., (Copyright © 2024 García-Llorca, Carta, Supuran and Eysteinsson.)

Mutations in the mouse microphthalmia-associated transcription factor ( Mitf ) gene affect retinal pigment epithelium (RPE) differentiation and development and can lead to hypopigmentation, microphthalmia, deafness, and blindness. For instance, an association has been established between loss-of-function mutations in the mouse Mitf gene and a variety of human retinal diseases, including Waardenburg type 2 and Tietz syndromes. Although there is evidence showing that mice with the homozygous Mitf mi mutation manifest microphthalmia and osteopetrosis, there are limited or no data on the effects of the heterozygous condition in the eye. Mitf mice can therefore be regarded as an important model system for the study of human disease. Thus, we characterized Mitf mi/+ mice at 1, 3, 12, and 18 months old in comparison with age-matched wild-type mice. The light- and dark-adapted electroretinogram (ERG) recordings showed progressive cone-rod dystrophy in Mitf mi/+ mice. The RPE response was reduced in the mutant in all age groups studied. Progressive loss of pigmentation was found in Mitf mi/+ mice. Histological retinal sections revealed evidence of retinal degeneration in Mitf mi/+ mice at older ages. For the first time, we report a mouse model of progressive cone-rod dystrophy and RPE dysfunction with a mutation in the Mitf gene., Competing Interests: The authors declare no conflicts of interest.

It is important to study the development of retinal vasculature in retinopathies in which abnormal vessel growth can ultimately lead to vision loss. Mutations in the microphthalmia-associated transcription factor (Mitf) gene show hypopigmentation, microphthalmia, retinal degeneration, and in some cases, blindness. In vivo imaging of the mouse retina by noninvasive means is vital for eye research. However, given its small size, mouse fundus imaging is difficult and might require specialized tools, maintenance, and training. In this study, we have developed a unique software enabling analysis of the retinal vessel diameter in mice with an automated program written in MATLAB. Fundus photographs were obtained with a commercial fundus camera system following an intraperitoneal injection of a fluorescein salt solution. Images were altered to enhance contrast, and the MATLAB program permitted extracting the mean vascular diameter automatically at a predefined distance from the optic disk. The vascular changes were examined in wild-type mice and mice with various mutations in the Mitf gene by analyzing the retinal vessel diameter. The custom-written MATLAB program developed here is practical, easy to use, and allows researchers to analyze the mean diameter and mean total diameter, as well as the number of vessels from the mouse retinal vasculature, conveniently and reliably.

It has been demonstrated previously that a variety of carbonic anhydrase inhibitors (CAIs) can induce vasodilation in pre-contracted retinal arteriolar segments although with different efficacy and potency. Since the CAIs tested so far are able to permeate cell membranes and inhibit both intracellular and extracellular isoforms of the enzyme, it is not clear whether extra- or intracellular isoforms or mechanisms are mediating their vasodilatory effects. By means of small wire myography, we have tested the effects of four new CAIs on wall tension in pre-contracted retinal arteriolar segments that demonstrably do not enter cell membranes but have high affinity to both cytosolic and membrane-bound isoforms of CA. At concentrations between 10 -6 M to 10 -3 M, none of the four membrane impermeant CAIs had any significant effect on arteriolar wall tension, while the membrane permeant CAI benzolamide (10 -3 M) fully dilated all arteriolar segments tested. This suggests that CAI act as vasodilators through cellular mechanisms located in the cytoplasm of vascular cells.

Purpose: Mice carrying pathogenic variants in the microphthalmia transcription factor (Mitf) gene show structural and functional changes in the retina and retinal pigment epithelium. The purpose of this study was to assess the vascular changes in Mitf mice carrying pathogenic variants by determining their retinal vessel diameter., Methods: Mice examined in this study were: B6-Mitf mi-vga9/+ (n = 6), B6-Mitf mi-enu22(398) /Mitf mi-enu22(398) (n = 6) and C57BL/6J wild type mice (n = 6), all 3 months old. Fundus images were taken with a Micron IV camera after intraperitoneal injection of fluorescein salt. Images were adjusted to enhance contrast and a custom written MATLAB program used to extract the mean vascular diameter at a pre-defined distance from the optic disc. The number of vessels, mean diameter and mean total diameter were examined., Results: The mean diameter of retinal veins in Mitf mi-enu22(398) /Mitf mi-enu22(398) mice was 18.8% larger than in wild type (p = 0.026). No differences in the mean diameter of the retinal arteries were found between the genotypes. Mitf mi-enu22(398) /Mitf mi-enu22(398) mice have 17.2% more retinal arteries (p = 0.026), and 15.6% more retinal veins (p = 0.041) than wild type. A 24.8% increase was observed in the mean combined arterial diameter in mice with the Mitf mi-enu22(398)/ Mitf mi-enu22(398) compared to wild type mice (p = 0.024). A 38.6% increase was found in the mean combined venular diameter in mice with the Mitf mi-enu22(398) /Mitf mi-enu22(398) pathogenic variation as compared to wild type (p = 0.004). The mean combined retinal venular diameter in the Mitf mi-vga9/+ mice was 17.8% larger than in wild type (p = 0.03)., Conclusion: An increase in vascularization of the retina in Mitf mi-enu22(398) /Mitf mi-enu22(398) mice was found, indicating an increased demand for blood flow to the retina., (© 2022 The Authors. Acta Ophthalmologica published by John Wiley & Sons Ltd on behalf of Acta Ophthalmologica Scandinavica Foundation.)

Purpose: In this study, we investigated the vasodilation properties on pre-contracted retinal arteries of a restricted series of carbonic anhydrase inhibitors (CAIs) of the sulfonamide type with enhanced lipophilicity, to assess if it affects the potency of CAIs as vasodilators., Methods: Carbonic anhydrase (CA) inhibition and in vitro kinetics of the compounds designed and synthesized for testing in this study were assessed by extracting human CA isoform proteins (hCA) from human cells expressing the isoforms of interest, and then measure the affinity of the novel compound for the hCAs by stopped-flow CO 2 hydrase spectroscopy. Lipophilicity of compounds was measured by obtaining their octanol-water partition coefficient, expressed as calculated logP. Porcine eyes were obtained from a local abattoir, and the wall tension of porcine retinal arteriole segments dissected from the eyes was measured with small wire vessel myography. The effects of the CA compounds on wall tension were assessed by adding them to the myography bath, after pre-contracting the vessel by prostaglandin analog U-46619., Results: All compounds induced vasodilation but at different concentrations. Among the tested compounds the most potent vasodilators were found to be the seleno-compound 4 and sulfur-ether compound 8 with EC 50 values of 7.13 × 10 -5 and 7.93 × 10 -5 M, respectively, whereas the remaining ones induced complete vasodilation at EC 50 comprised within the sub millimolar range., Conclusions: All the data reported in this study (i.e. results from myography, in vitro kinetics and LogPs) confirm the important role played by several CA isoforms in vasodilation, although the precise mechanism of action still remains to be elucidated.