Extracellular lactate as an alternative energy source for retinal bipolar cells.

Retinal bipolar and amacrine cells receive visual information from photoreceptors and participate in the first steps of image processing in the retina. Several studies have suggested the operation of aerobic glycolysis and a lactate shuttle system in the retina due to the high production of this metabolite under aerobic conditions. However, whether bipolar cells form part of this metabolic circuit remains unclear. Here, we show that the monocarboxylate transporter 2 is expressed and functional in inner retinal neurons. Additionally, we used genetically encoded FRET nanosensors to demonstrate the ability of inner retinal neurons to consume extracellular lactate as an alternative to glucose. In rod bipolar cells, lactate consumption allowed cells to maintain the homeostasis of ions and electrical responses. We also found that lactate synthesis and transporter inhibition caused functional alterations and an increased rate of cell death. Overall, our data shed light on a notable but still poorly understood aspect of retinal metabolism.

Immune response induced by coinfection of the sea louse Caligus rogercresseyi and the intracellular bacteria Piscirickettsia salmonis in vaccinated Atlantic salmon.

Recently, we showed that Atlantic salmon vaccinated against Piscirickettsia salmonis lose their protection upon coinfection with Caligus rogercresseyi (sea lice). However, the causes of the overriding effect of C. rogercresseyi infection have not been elucidated, and the molecular basis of the cellular and humoral immune responses upon C. rogercresseyi infection has not been described for vaccinated salmon. Therefore, we studied changes in the transcription of immune genes in vaccinated Atlantic salmon that were experimentally challenged by co-infecting them with C. rogercresseyi and P. salmonis. In general, coinfection treatments showed immune gene expression similar to treatments with a single P. salmonis infection, showing a decreased cellular response. However, a high variance was found between individual fish in the case of crucial cellular immune genes, with a few fish reacting overwhelmingly highly compared to the majority. This supports our previous findings on vaccination response variation and reinforces the idea that vaccination failures in the field might be caused by an overwhelming amount of vaccinated fish that display a deficient immune response to the infection.

Evaluation of Photobiomodulation and Boldine as Alternative Treatment Options in Two Diabetic Retinopathy Models.

Diabetic retinopathy causes progressive and irreversible damage to the retina through activation of inflammatory processes, overproduction of oxidative species, and glial reactivity, leading to changes in neuronal function and finally ischemia, edema, and hemorrhages. Current treatments are invasive and mostly applied at advanced stages, stressing the need for alternatives. To this end, we tested two unconventional and potentially complementary non-invasive treatment options: Photobiomodulation, the stimulation with near-infrared light, has shown promising results in ameliorating retinal pathologies and insults in several studies but remains controversial. Boldine, on the other hand, is a potent natural antioxidant and potentially useful to prevent free radical-induced oxidative stress. To establish a baseline, we first evaluated the effects of diabetic conditions on the retina with immunofluorescence, histological, and ultrastructural analysis in two diabetes model systems, obese LepRdb/db mice and organotypic retinal explants, and then tested the potential benefits of photobiomodulation and boldine treatment in vitro on retinal explants subjected to high glucose concentrations, mimicking diabetic conditions. Our results suggest that the principal subcellular structures affected by these conditions were mitochondria in the inner segment of photoreceptors, which displayed morphological changes in both model systems. In retinal explants, lactate metabolism, assayed as an indicator of mitochondrial function, was altered, and decreased photoreceptor viability was observed, presumably as a consequence of increased oxidative-nitrosative stress. The latter was reduced by boldine treatment in vitro, while photobiomodulation improved mitochondrial metabolism but was insufficient to prevent retinal structural damage caused by high glucose. These results warrant further research into alternative and complementary treatment options for diabetic retinopathy.

Imaging of lactate metabolism in retinal Müller cells with a FRET nanosensor.

Müller cells, the glial cells of the retina, provide metabolic support for photoreceptors and inner retinal neurons, and have been proposed as source of the significant lactate production of this tissue. To better understand the role of lactate in retinal metabolism, we expressed a lactate and a glucose nanosensor in organotypic mouse retinal explants cultured for 14 days, and used FRET imaging in acute vibratome sections of the explants to study metabolite flux in real time. Pharmacological manipulation with specific monocarboxylate transporter (MCT) inhibitors and immunohistochemistry revealed the functional expression of MCT1, MCT2 and MCT4 in Müller cells of retinal explants. The introduction of FRET nanosensors to measure key metabolites at the cellular level may contribute to a better understanding of heretofore poorly understood issues in retinal metabolism.

Determination of the Severity of Pulpitis by Immunohistological Analysis and Comparison with the Clinical Picture.

INTRODUCTION: Inflammation of the dental pulp due to caries is a highly prevalent pathology which causes intense pain. Here, we sought to correlate the clinical picture with the histopathology of the affected tissue. The interaction between nociceptive neurons and immune cells is fundamental to regulate the inflammatory response, but little is known about the glial network involved in this process, and its impact on caries pathogenesis. METHODS: This study characterized Schwann cells and other neuroimmune components in human dental pulps with reversible and symptomatic irreversible pulpitis (IP). Twenty eight human teeth were extracted for reasons beyond the scope of this study. Twelve were diagnosed as reversible and symptomatic IP respectively, and 4 as controls. The teeth were decalcified, processed for immunolabeling and analyzed with confocal microscopy. RESULTS: Symptomatic IP was characterized by a significantly higher density of neutrophils, and the release of neutrophil extracellular traps. Between IP and healthy controls, there were significant differences in the density of Schwann cells, macrophages, and neutrophils, in addition to morphological alterations. In IP, Schwann cell arborization extended toward the pulpodentinal interface along with more spindle-shaped cell bodies, while some macrophages displayed a distinct fusiform phenotype. CONCLUSIONS: The dental pulp has a complex multicellular organization and its pulpodentinal interface acts as a barrier in which Schwann and immune cells are distributed strategically to stop the progress of pathogens. A synergistic interaction of Schwann cells with immune cells creates a novel perspective to better understand the role of these glial cells and their active participation in pulpal inflammation.

The interplay between α7 nicotinic acetylcholine receptors, pannexin-1 channels and P2X7 receptors elicit exocytosis in chromaffin cells.

Pannexin-1 (Panx1) forms plasma membrane channels that allow the exchange of small molecules between the intracellular and extracellular compartments, and are involved in diverse physiological and pathological responses in the nervous system. However, the signaling mechanisms that induce their opening still remain elusive. Here, we propose a new mechanism for Panx1 channel activation through a functional crosstalk with the highly Ca2+ permeable α7 nicotinic acetylcholine receptor (nAChR). Consistent with this hypothesis, we found that activation of α7 nAChRs induces Panx1-mediated dye uptake and ATP release in the neuroblastoma cell line SH-SY5Y-α7. Using membrane permeant Ca2+ chelators, total internal reflection fluorescence microscopy in SH-SY5Y-α7 cells expressing a membrane-tethered GCAMP3, and Src kinase inhibitors, we further demonstrated that Panx1 channel opening depends on Ca2+ signals localized in submembrane areas, as well as on Src kinases. In turn, Panx1 channels amplify cytosolic Ca2+ signals induced by the activation of α7 nAChRs, by a mechanism that seems to involve ATP release and P2X7 receptor activation, as hydrolysis of extracellular ATP with apyrase or blockage of P2X7 receptors with oxidized ATP significantly reduces the α7 nAChR-Ca2+ signal. The physiological relevance of this crosstalk was also demonstrated in neuroendocrine chromaffin cells, wherein Panx1 channels and P2X7 receptors contribute to the exocytotic release of catecholamines triggered by α7 nAChRs, as measured by amperometry. Together these findings point to a functional coupling between α7 nAChRs, Panx1 channels and P2X7 receptors with physiological relevance in neurosecretion.

Physiological assessment of high glucose neurotoxicity in mouse and rat retinal explants.

One of the tissues of the central nervous system most affected by diabetes is the retina. Despite a growing understanding of the biochemical processes involved in glucose toxicity, little is known about the physiological consequences of chronic high glucose (HG) on individual neurons and neuronal circuits. Electroretinogram recordings suggest that retinal bipolar cells (BCs), which filter and transmit photoreceptor output to the inner retina, are among the first cells affected by diabetic conditions, and may therefore serve as sensitive early biomarkers for incipient neuronal damage caused in diabetes. Here, we comparatively assessed retinal integrity, calcium responses, and the electrophysiological profiles of specific BC types of mouse and rat organotypic retinal explants after 1 to 3 weeks in tissue culture, under moderate glucose (MG) and HG conditions. While the retinal layers of both rodent species displayed a progressively reduced thickness in culture, BCs retained their electrophysiological profiles and remained morphologically identifiable for up to 2 weeks. Responses to glutamate and endogenous inhibitory responses were routinely observed, indicating that the retinal circuitry remained intact during this period. Significant physiological differences between MG and HG conditions were evident in calcium signals and in the time course of responses to glutamate, but the voltage-gated current profiles of BCs displayed only minor variations. Overall, rat retina appeared slightly more sensitive to HG levels compared with mouse. In conclusion, electrophysiological analysis of neuronal function in rodent retinal explants is useful for the study of early damage due to HG neurotoxicity.

Impact of stress and mitigating information on evaluations, attributions, affect, disciplinary choices, and expectations of compliance in mothers at high and low risk for child physical abuse.

The objective is to know if high-risk mothers for child physical abuse differ in their evaluations, attributions, negative affect, disciplinary choices for children's behavior, and expectations of compliance. The effect of a stressor and the introduction of mitigating information are analyzed. Forty-seven high-risk and 48 matched low-risk mothers participated in the study. Mothers' information processing and disciplinary choices were examined using six vignettes depicting a child engaging in different transgressions. A four-factor design with repeated measures on the last two factors was used. High-risk mothers reported more hostile intent, global and internal attributions, more use of power assertion discipline, and less induction. A risk group by child transgression interaction and a risk group by mitigating information interaction were found. Results support the social information-processing model of child physical abuse, which suggests that high-risk mothers process child-related information differently and use more power assertive and less inductive disciplinary techniques.