Preservation of Intrinsically Photosensitive Retinal Ganglion Cells (ipRGCs) in Late Adult Mice: Implications as a Potential Biomarker for Early Onset Ocular Degenerative Diseases.

Purpose: Intrinsically photosensitive retinal ganglion cells (ipRGCs) play a crucial role in non-image-forming visual functions. Given their significant loss observed in various ocular degenerative diseases at early stages, this study aimed to assess changes in both the morphology and associated behavioral functions of ipRGCs in mice between 6 (mature) and 12 (late adult) months old. The findings contribute to understanding the preservation of ipRGCs in late adults and their potential as a biomarker for early ocular degenerative diseases. Methods: Female and male C57BL/6J mice were used to assess the behavioral consequences of aging to mature and old adults, including pupillary light reflex, light aversion, visual acuity, and contrast sensitivity. Immunohistochemistry on retinal wholemounts from these mice was then conducted to evaluate ipRGC dendritic morphology in the ganglion cell layer (GCL) and inner nuclear layer (INL). Results: Morphological analysis showed that ipRGC dendritic field complexity was remarkably stable through 12 months old of age. Similarly, the pupillary light reflex, visual acuity, and contrast sensitivity were stable in mature and old adults. Although alterations were observed in ipRGC-independent light aversion distinct from the pupillary light reflex, aged wild-type mice continuously showed enhanced light aversion with dilation. No effect of sex was observed in any tests. Conclusions: The preservation of both ipRGC morphology and function highlights the potential of ipRGC-mediated function as a valuable biomarker for ocular diseases characterized by early ipRGC loss. The consistent stability of ipRGCs in mature and old adult mice suggests that detected changes in ipRGC-mediated functions could serve as early indicators or diagnostic tools for early-onset conditions such as Alzheimer's disease, Parkinson's disease, and diabetes, where ipRGC loss has been documented.

PTEN Expression Regulates Gap Junction Connectivity in the Retina.

Manipulation of the phosphatase and tensin homolog (PTEN) pathway has been suggested as a therapeutic approach to treat or prevent vision loss due to retinal disease. In this study, we investigated the effects of deleting one copy of Pten in a well-characterized class of retinal ganglion cells called α-ganglion cells in the mouse retina. In Pten +/- retinas, α-ganglion cells did not exhibit major changes in their dendritic structure, although most cells developed a few, unusual loop-forming dendrites. By contrast, α-ganglion cells exhibited a significant decrease in heterologous and homologous gap junction mediated cell coupling with other retinal ganglion and amacrine cells. Additionally, the majority of OFF α-ganglion cells (12/18 cells) formed novel coupling to displaced amacrine cells. The number of connexin36 puncta, the predominant connexin that mediates gap junction communication at electrical synapses, was decreased by at least 50% on OFF α-ganglion cells. Reduced and incorrect gap junction connectivity of α-ganglion cells will affect their functional properties and alter visual image processing in the retina. The anomalous connectivity of retinal ganglion cells would potentially limit future therapeutic approaches involving manipulation of the Pten pathway for treating ganglion cell degeneration in diseases like glaucoma, traumatic brain injury, Parkinson's, and Alzheimer's diseases.

Multiple cell types form the VIP amacrine cell population.

Amacrine cells are a heterogeneous group of interneurons that form microcircuits with bipolar, amacrine and ganglion cells to process visual information in the inner retina. This study has characterized the morphology, neurochemistry and major cell types of a VIP-ires-Cre amacrine cell population. VIP-tdTomato and -Confetti (Brainbow2.1) mouse lines were generated by crossing a VIP-ires-Cre line with either a Cre-dependent tdTomato or Brainbow2.1 reporter line. Retinal sections and whole-mounts were evaluated by quantitative, immunohistochemical, and intracellular labeling approaches. The majority of tdTomato and Confetti fluorescent cell bodies were in the inner nuclear layer (INL) and a few cell bodies were in the ganglion cell layer (GCL). Fluorescent processes ramified in strata 1, 3, 4, and 5 of the inner plexiform layer (IPL). All tdTomato fluorescent cells expressed syntaxin 1A and GABA-immunoreactivity indicating they were amacrine cells. The average VIP-tdTomato fluorescent cell density in the INL and GCL was 535 and 24 cells/mm2 , respectively. TdTomato fluorescent cells in the INL and GCL contained VIP-immunoreactivity. The VIP-ires-Cre amacrine cell types were identified in VIP-Brainbow2.1 retinas or by intracellular labeling in VIP-tdTomato retinas. VIP-1 amacrine cells are bistratified, wide-field cells that ramify in strata 1, 4, and 5, VIP-2A and 2B amacrine cells are medium-field cells that mainly ramify in strata 3 and 4, and VIP-3 displaced amacrine cells are medium-field cells that ramify in strata 4 and 5 of the IPL. VIP-ires-Cre amacrine cells form a neuropeptide-expressing cell population with multiple cell types, which are likely to have distinct roles in visual processing.

Percepção de enfermeiros sobre o processo de gestão em um hospital universitário / Perception of nurses on the management process in a university hospital

Objetivo: conhecer a percepção de enfermeiros gerentes sobre o processo de gestão na enfermagem.Método: estudo descritivo com abordagem qualitativa, realizado em um hospital universitário do Sul doBrasil, com seis enfermeiros gerentes. Os dados foram coletados por meio de entrevista semiestruturada,transcritas e submetidas à Técnica de Análise de Conteúdo na modalidade Análise Categorial. Resultados:emergiram três categorias >, > e >. Conclusão: os enfermeiros em cargo de gerência de umhospital universitário do Sul do Brasil percebem o processo de gestão sob três principais aspectos:competências gerenciais e funções do enfermeiro gerente, gerência compartilhada e gerenciamento docuidado.

Prox1 Is a Marker for AII Amacrine Cells in the Mouse Retina.

The transcription factor Prox1 is expressed in multiple cells in the retina during eye development. This study has focused on neuronal Prox1 expression in the inner nuclear layer (INL) of the adult mouse retina. Prox1 immunostaining was evaluated in vertical retinal sections and whole mount preparations using a specific antibody directed to the C-terminus of Prox1. Strong immunostaining was observed in numerous amacrine cell bodies and in all horizontal cell bodies in the proximal and distal INL, respectively. Some bipolar cells were also weakly immunostained. Prox1-immunoreactive amacrine cells expressed glycine, and they formed 35 ± 3% of all glycinergic amacrine cells. Intracellular Neurobiotin injections into AII amacrine cells showed that all gap junction-coupled AII amacrine cells express Prox1, and no other Prox1-immunostained amacrine cells were in the immediate area surrounding the injected AII amacrine cell. Prox1-immunoreactive amacrine cell bodies were distributed across the retina, with their highest density (3887 ± 160 cells/mm2) in the central retina, 0.5 mm from the optic nerve head, and their lowest density (3133 ± 350 cells/mm2) in the mid-peripheral retina, 2 mm from the optic nerve head. Prox1-immunoreactive amacrine cell bodies comprised ~9.8% of the total amacrine cell population, and they formed a non-random mosaic with a regularity index (RI) of 3.4, similar to AII amacrine cells in the retinas of other mammals. Together, these findings indicate that AII amacrine cells are the predominant and likely only amacrine cell type strongly expressing Prox1 in the adult mouse retina, and establish Prox1 as a marker of AII amacrine cells.

Peripheral Sensory Neurons Expressing Melanopsin Respond to Light.

The ability of light to cause pain is paradoxical. The retina detects light but is devoid of nociceptors while the trigeminal sensory ganglia (TG) contain nociceptors but not photoreceptors. Melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs) are thought to mediate light-induced pain but recent evidence raises the possibility of an alternative light responsive pathway independent of the retina and optic nerve. Here, we show that melanopsin is expressed in both human and mouse TG neurons. In mice, they represent 3% of small TG neurons that are preferentially localized in the ophthalmic branch of the trigeminal nerve and are likely nociceptive C fibers and high-threshold mechanoreceptor Aδ fibers based on a strong size-function association. These isolated neurons respond to blue light stimuli with a delayed onset and sustained firing, similar to the melanopsin-dependent intrinsic photosensitivity observed in ipRGCs. Mice with severe bilateral optic nerve crush exhibit no light-induced responses including behavioral light aversion until treated with nitroglycerin, an inducer of migraine in people and migraine-like symptoms in mice. With nitroglycerin, these same mice with optic nerve crush exhibit significant light aversion. Furthermore, this retained light aversion remains dependent on melanopsin-expressing neurons. Our results demonstrate a novel light-responsive neural function independent of the optic nerve that may originate in the peripheral nervous system to provide the first direct mechanism for an alternative light detection pathway that influences motivated behavior.

Targeted Deletion of Vesicular GABA Transporter from Retinal Horizontal Cells Eliminates Feedback Modulation of Photoreceptor Calcium Channels.

The cellular mechanisms underlying feedback signaling from horizontal cells to photoreceptors, which are important for the formation of receptive field surrounds of early visual neurons, remain unsettled. Mammalian horizontal cells express a complement of synaptic proteins that are necessary and sufficient for calcium-dependent exocytosis of inhibitory neurotransmitters at their contacts with photoreceptor terminals, suggesting that they are capable of releasing GABA via vesicular release. To test whether horizontal cell vesicular release is involved in feedback signaling, we perturbed inhibitory neurotransmission in these cells by targeted deletion of the vesicular GABA transporter (VGAT), the protein responsible for the uptake of inhibitory transmitter by synaptic vesicles. To manipulate horizontal cells selectively, an iCre mouse line with Cre recombinase expression controlled by connexin57 (Cx57) regulatory elements was generated. In Cx57-iCre mouse retina, only horizontal cells expressed Cre protein, and its expression occurred in all retinal regions. After crossing with a VGAT(flox/flox) mouse line, VGAT was selectively eliminated from horizontal cells, which was confirmed immunohistochemically. Voltage-gated ion channel currents in horizontal cells of Cx57-VGAT(-/-) mice were the same as Cx57-VGAT(+/+) controls, as were the cell responses to the ionotropic glutamate receptor agonist kainate, but the response to the GABAA receptor agonist muscimol in Cx57-VGAT(-/-) mice was larger. In contrast, the feedback inhibition of photoreceptor calcium channels, which in control animals is induced by horizontal cell depolarization, was completely absent in Cx57-VGAT(-/-) mice. The results suggest that vesicular release of GABA from horizontal cells is required for feedback inhibition of photoreceptors.

Hospitalização em unidade de terapia intensiva na voz de idosos e familiares / Hospitalization in intensive care units in the voice of the elderly and their families

Este estudo objetivou compreender as vivências do processo de hospitalização em uma unidade de terapia intensiva na voz de idosos e seus familiares. É uma pesquisa descritiva com abordagem qualitativa realizada em um hospital público do Rio Grande do Sul, do qual participaram seis idosos e oito familiares. A análise seguiu os passos da análise de conteúdo. A partir das informações, foi possível construir duas categorias: unidade de terapia intensiva como espaço desconhecido e necessário para manter a vida, e orientações da equipe de saúde que contribuam para enfrentar a internação em unidade de terapia intensiva. Entende-se que discutir os aspectos relativos à hospitalização de idosos contribui para qualificar a assistência de enfermagem nesse espaço, isto pelas dificuldades enfrentadas para assistir de forma integral essa população, necessitando de abordagem própria e que ultrapasse o campo individual e curativo. Também, destaca-se que, no cuidado ao idoso hospitalizado em unidade de terapia intensiva, devem ser incluídas as necessidades da família uma vez que ela precisa de informações claras sobre as condições clínicas de seu familiar, além de apoio para o enfrentamento das dificuldades decorrentes da hospitalização. (AU)

This study aimed to understand the experiences of the process of hospitalization in an intensive care unit in the voice of older people and their families. It is a descriptive research with a qualitative approachperformed in a public hospital in Rio Grande do Sul in which six elderly and eight family members participated. The analysis followed the steps of content analysis. From the information, it was possible to form two categories: intensive care unit as unknown space and needed to maintain life, and guidelines of health staff who contribute to confront the hospitalization in intensive care unit. It is understood that discuss the aspects relating to hospitalization of elderly contributes to qualify for nursing care in this area. This by difficulties face to assist in an integral way this population, requiring its own approach and that exceeds the individual field and dressing. Also, it is noteworthy that in the care of elderly patients hospitalized in the intensive care unit should be included the needs of the family, since it needs clear information on the clinical conditions of his family, as well as support for coping with the difficulties arising from the hospitalization. (AU)

Hospitalização em unidade de terapia intensiva na voz de idosos e familiares / Hospitalization in intensive care units in the voice of the elderly and their families

Este estudo objetivou compreender as vivências do processo de hospitalização em uma unidade de terapia intensiva na voz de idosos e seus familiares. É uma pesquisa descritiva com abordagem qualitativa realizada em um hospital público do Rio Grande do Sul, do qual participaram seis idosos e oito familiares. A análise seguiu os passos da análise de conteúdo. A partir das informações, foi possível construir duas categorias: unidade de terapia intensiva como espaço desconhecido e necessário para manter a vida, e orientações da equipe de saúde que contribuam para enfrentar a internação em unidade de terapia intensiva. Entende-se que discutir os aspectos relativos à hospitalização de idosos contribui para qualificar a assistência de enfermagem nesse espaço, isto pelas dificuldades enfrentadas para assistir de forma integral essa população, necessitando de abordagem própria e que ultrapasse o campo individual e curativo. Também, destaca-se que, no cuidado ao idoso hospitalizado em unidade de terapia intensiva, devem ser incluídas as necessidades da família uma vez que ela precisa de informações claras sobre as condições clínicas de seu familiar, além de apoio para o enfrentamento das dificuldades decorrentes da hospitalização.

This study aimed to understand the experiences of the process of hospitalization in an intensive care unit in the voice of older people and their families. It is a descriptive research with a qualitative approachperformed in a public hospital in Rio Grande do Sul in which six elderly and eight family members participated. The analysis followed the steps of content analysis. From the information, it was possible to form two categories: intensive care unit as unknown space and needed to maintain life, and guidelines of health staff who contribute to confront the hospitalization in intensive care unit. It is understood that discuss the aspects relating to hospitalization of elderly contributes to qualify for nursing care in this area. This by difficulties face to assist in an integral way this population, requiring its own approach and that exceeds the individual field and dressing. Also, it is noteworthy that in the care of elderly patients hospitalized in the intensive care unit should be included the needs of the family, since it needs clear information on the clinical conditions of his family, as well as support for coping with the difficulties arising from the hospitalization.

How do peri-implant mucositis and gingivitis respond to supragingival biofilm control - an intra-individual longitudinal cohort study.

PURPOSE: This single-arm study to compare the gingival with peri-implant mucosal inflammatory response to a mechanical supragingival-supramucosal biofilm control program. MATERIALS AND METHODS: Twenty-two participants (55.7 ± 11.2 years) with both gingivitis and periimplant mucositis were examined at days 0, 30 and 390 (full mouth/6 sites per tooth/implant [TTH/IMPL]) for visible plaque (VPI), gingival bleeding (GBI), modified plaque (mPlI) and bleeding indexes (mBI), probing depth (PD) and bleeding on probing (BOP). The biofilm control was carried out weekly in the first month and every 3 months thereafter. An intention-to-treat analysis was performed (drop-out rate = 8) and linear models were used against comparisons in order to look at the clustering of TTH/IMPL by each individual. RESULTS: VPI/mPlI and GBI/mBI reduced from day 0 onwards. Intra-group reductions (P < 0.05) were observed at day 30. PD values (in mm) were higher (P < 0.001) for IMPL than for TTH [mean difference (95% CI) at day 0: -1.10 (-1.58 to -0.63); day 30: -0.88 (-1.28 to -0.48); and day 390: -0.60 (-0.84 to -0.33)], where both groups showed reductions (P < 0.05) throughout the study. BOP was greater (P = 0.00001) for IMPL at baseline [mean difference (95% CI): -0.24 (-0.31 to -0.17)] but reduced (P = 0.00001) and showed similar levels to TTH from day 30 onwards. With regard to sites with the greatest PD, BOP reduced (P < 0.05) in both IMPL and TTH, with greater PD reductions observed for IMPL (P = 0.00001). CONCLUSIONS: The supragingival-supramucosal biofilm control benefited both teeth and implants.

Expression and cellular localization of the voltage-gated calcium channel α2δ3 in the rodent retina.

High-voltage-activated calcium channels are hetero-oligomeric protein complexes that mediate multiple cellular processes, including the influx of extracellular Ca(2+), neurotransmitter release, gene transcription, and synaptic plasticity. These channels consist of a primary α(1) pore-forming subunit, which is associated with an extracellular α(2)δ subunit and an intracellular ß auxiliary subunit, which alter the gating properties and trafficking of the calcium channel. The cellular localization of the α(2)δ(3) subunit in the mouse and rat retina is unknown. In this study using RT-PCR, a single band at ∼ 305 bp corresponding to the predicted size of the α(2)δ(3) subunit fragment was found in mouse and rat retina and brain homogenates. Western blotting of rodent retina and brain homogenates showed a single 123-kDa band. Immunohistochemistry with an affinity-purified antibody to the α(2)δ(3) subunit revealed immunoreactive cell bodies in the ganglion cell layer and inner nuclear layer and immunoreactive processes in the inner plexiform layer and the outer plexiform layer. α(2)δ(3) immunoreactivity was localized to multiple cell types, including ganglion, amacrine, and bipolar cells and photoreceptors, but not horizontal cells. The expression of the α(2)δ(3) calcium channel subunit to multiple cell types suggests that this subunit participates widely in Ca-channel-mediated signaling in the retina.

Immunohistochemical and calcium imaging methods in wholemount rat retina.

In this paper we describe the tools, reagents, and the practical steps that are needed for: 1) successful preparation of wholemount retinas for immunohistochemistry and, 2) calcium imaging for the study of voltage gated calcium channel (VGCC) mediated calcium signaling in retinal ganglion cells. The calcium imaging method we describe circumvents issues concerning non-specific loading of displaced amacrine cells in the ganglion cell layer.

Qualificação da equipe de enfermagem mediante pesquisa convergente assistencial: contribuições ao cuidado do idoso hospitalizado / Qualification of the nursing team by means of convergent-care research: contributions to the care of the hospitalized elderly person

Objetivo: Analisar as contribuições da prática educativa na qualificação da equipe de enfermagem, para prestar o cuidado a idosos hospitalizados. Método: Estudo qualitativo, descritivo que utilizou os preceitos teórico-metodológicos da pesquisa convergente assistencial, para produção e análise das informações. Participaram vinte profissionais de enfermagem, que trabalham em uma unidade de internação de clínica médica. Resultados: Para a maioria dos sujeitos, o idoso requer cuidados diferenciados e a prática educativa é uma ferramenta que pode ser utilizada para contribuir na qualificação dos integrantes da equipe de enfermagem. Tal aprimoramento, melhora a atenção de enfermagem ao idoso hospitalizado. Conclusão: Conclui-se que a prática educativa constitui-se em instrumento de qualificação e atualização para os integrantes da equipe de enfermagem.

Loss of outer retinal neurons and circuitry alterations in the DBA/2J mouse.

PURPOSE: The DBA/2J mouse line develops essential iris atrophy, pigment dispersion, and glaucomatous age-related changes, including an increase of IOP, optic nerve atrophy, and retinal ganglion cell (RGC) death. The aim of this study was to evaluate possible morphological changes in the outer retina of the DBA/2J mouse concomitant with disease progression and aging, based on the reduction of both the a- and b-waves and photopic flicker ERGs in this mouse line. METHODS: Vertically sectioned DBA/2J mice retinas were evaluated at 3, 8, and 16 months of age using photoreceptor, horizontal, and bipolar cell markers. Sixteen-month-old C57BL/6 mice retinas were used as controls. RESULTS: The DBA/2J mice had outer retinal degeneration at all ages, with the most severe degeneration in the oldest retinas. At 3 months of age, the number of photoreceptor cells and the thickness of the OPL were reduced. In addition, there was a loss of horizontal and ON-bipolar cell processes. At 8 months of age, RGC degeneration occurred in patches, and in the outer retina overlying these patches, cone morphology was impaired with a reduction in size as well as loss of outer segments and growth of horizontal and bipolar cell processes into the outer nuclear layer. At 16 months of age, connectivity between photoreceptors and horizontal and bipolar cell processes overlying these patches was lost. CONCLUSIONS: Retinal degeneration in DBA/2J mice includes photoreceptor death, loss of bipolar and horizontal cell processes, and loss of synaptic contacts in an aging-dependent manner.

Percepções de professores enfermeiros sobre a intersecção do trabalho assistencial e docente / Percepciones de profesores enfermeros sobre la intersección del trabajo asistencial y docente / Perceptions of nurse professors about the intersection of assistantial and teaching work

Objetivo: conhecer as percepções de professores enfermeiros sobre a importância da experiência assistencial em relação ao exercício do trabalho docente. Método: a pesquisa caracteriza-se como um estudo exploratório-descritivo, com abordagem qualitativa. Na apreciação dos dados, utilizou-se a análise de conteúdo. Os participantes da pesquisa foram sete professores universitários que, anteriormente à prática docente haviam tido experiência na assistência. Resultados: o estudo mostra que as experiências no campo da prática assistencial contribuem para o desenvolvimento de conteúdos, sobretudo no que se refere à intersecção entre teoria e prática. Conclusão: a experiência na assistência contribui, no que se refere ao ensino, em relacionar teoria e prática, contudo, isso não se evidencia em outras atividades como pesquisa e extensão.

Aim: to understand the perceptions of nurse professors about the importance of healthcare experience regarding to the exercise of teaching. Method: The study is characterized as a descriptive exploratory study with a qualitative approach. In evaluating the data we used content analysis. Survey participants were seven university professors prior teaching practice had experience in assisting. Results: The study shows that the experience in the field of health care practice, contribute to the development of content, especially as regards to the intersection between theory and practice, Conclusion: The experience in assisting contributes with regard to education, to relate theory and practice, however, this is not evident in other activities such research an extension.

Objetivo: conocer las percepciones de los profesores enfermeros sobre la importancia de la experiencia asistencial en relación al ejercicio de la docencia. Método: La investigación se caracteriza como estudio exploratorio descriptivo con enfoque cualitativo. En la evaluación de los datos, se utilizó el análisis de contenido. Los participantes de la investigación fueron siete profesores universitarios que antes de la práctica docente habían tenido experiencia en la asistencia. Resultados: El estudio muestra que las experiencias en el campo de la práctica asistencial contribuyen al desarrollo de los contenidos, especialmente en lo que se refiere a la intersección entre teoría y práctica. Conclusión: La experiencia en la asistencia contribuye, con respecto a la enseñanza, a relacionar teoría y práctica, sin embargo, esto no se demuestra en otras actividades como investigación o extensión.

Melanopsin ganglion cells are the most resistant retinal ganglion cell type to axonal injury in the rat retina.

We report that the most common retinal ganglion cell type that remains after optic nerve transection is the M1 melanopsin ganglion cell. M1 ganglion cells are members of the intrinsically photosensitive retinal ganglion cell population that mediates non-image-forming vision, comprising ∼2.5% of all ganglion cells in the rat retina. In the present study, M1 ganglion cells comprised 1.7±1%, 28±14%, 55±13% and 82±8% of the surviving ganglion cells 7, 14, 21 and 60 days after optic nerve transection, respectively. Average M1 ganglion cell somal diameter and overall morphological appearance remained unchanged in non-injured and injured retinas, suggesting a lack of injury-induced degeneration. Average M1 dendritic field size increased at 7 and 60 days following optic nerve transection, while average dendritic field size remained similar in non-injured retinas and in retinas at 14 and 21 days after optic nerve transection. These findings demonstrate that M1 ganglion cells are more resistant to injury than other ganglion cell types following optic nerve injury, and provide an opportunity to develop pharmacological or genetic therapeutic approaches to mitigate ganglion cell death and save vision following optic nerve injury.

The RNA binding protein RBPMS is a selective marker of ganglion cells in the mammalian retina.

There are few neurochemical markers that reliably identify retinal ganglion cells (RGCs), which are a heterogeneous population of cells that integrate and transmit the visual signal from the retina to the central visual nuclei. We have developed and characterized a new set of affinity-purified guinea pig and rabbit antibodies against RNA-binding protein with multiple splicing (RBPMS). On western blots these antibodies recognize a single band at 〜24 kDa, corresponding to RBPMS, and they strongly label RGC and displaced RGC (dRGC) somata in mouse, rat, guinea pig, rabbit, and monkey retina. RBPMS-immunoreactive cells and RGCs identified by other techniques have a similar range of somal diameters and areas. The density of RBPMS cells in mouse and rat retina is comparable to earlier semiquantitative estimates of RGCs. RBPMS is mainly expressed in medium and large DAPI-, DRAQ5-, NeuroTrace- and NeuN-stained cells in the ganglion cell layer (GCL), and RBPMS is not expressed in syntaxin (HPC-1)-immunoreactive cells in the inner nuclear layer (INL) and GCL, consistent with their identity as RGCs, and not displaced amacrine cells. In mouse and rat retina, most RBPMS cells are lost following optic nerve crush or transection at 3 weeks, and all Brn3a-, SMI-32-, and melanopsin-immunoreactive RGCs also express RBPMS immunoreactivity. RBPMS immunoreactivity is localized to cyan fluorescent protein (CFP)-fluorescent RGCs in the B6.Cg-Tg(Thy1-CFP)23Jrs/J mouse line. These findings show that antibodies against RBPMS are robust reagents that exclusively identify RGCs and dRGCs in multiple mammalian species, and they will be especially useful for quantification of RGCs.

Expression of voltage-gated calcium channel α(2)δ(4) subunits in the mouse and rat retina.

High-voltage activated Ca channels participate in multiple cellular functions, including transmitter release, excitation, and gene transcription. Ca channels are heteromeric proteins consisting of a pore-forming α(1) subunit and auxiliary α(2)δ and ß subunits. Although there are reports of α(2)δ(4) subunit mRNA in the mouse retina and localization of the α(2)δ(4) subunit immunoreactivity to salamander photoreceptor terminals, there is a limited overall understanding of its expression and localization in the retina. α(2)δ(4) subunit expression and distribution in the mouse and rat retina were evaluated by using reverse transcriptase polymerase chain reaction, western blot, and immunohistochemistry with specific primers and a well-characterized antibody to the α(2)δ(4) subunit. α(2)δ(4) subunit mRNA and protein are present in mouse and rat retina, brain, and liver homogenates. Immunostaining for the α(2)δ(4) subunit is mainly localized to Müller cell processes and endfeet, photoreceptor terminals, and photoreceptor outer segments. This subunit is also expressed in a few displaced ganglion cells and bipolar cell dendrites. These findings suggest that the α(2)δ(4) subunit participates in the modulation of L-type Ca(2+) current regulating neurotransmitter release from photoreceptor terminals and Ca(2+)-dependent signaling pathways in bipolar and Müller cells.

Tracer coupling of intrinsically photosensitive retinal ganglion cells to amacrine cells in the mouse retina.

Intrinsically photosensitive retinal ganglion cells (ipRGCs) are a subtype of ganglion cell in the mammalian retina that expresses the photopigment melanopsin and drives non-image-forming visual functions. Three morphological subtypes of ipRGCs (M1, M2, and M3) have been described based on their dendritic stratifications in the inner plexiform layer (IPL), but the question of their potential interactions via electrical coupling remains unsettled. In this study, we have addressed this question in the mouse retina by, injecting the tracer Neurobiotin into ipRGCs that had been genetically labelled with the fluorescent protein, tdTomato. We confirmed the presence of the M1-M3 subtypes of ipRGCs based on their distinct dendritic stratifications. All three subtypes were tracer coupled to putative amacrine cells situated within the ganglion cell layer (GCL) but not the inner nuclear layer (INL). The cells tracer coupled to the M1 and M2 cells were shown to be widefield GABA-immunoreactive amacrine cells. We found no evidence of homologous tracer coupling of ipRGCs or heterologous coupling to other types of ganglion cells.

Neuronal connexin-36 can functionally replace connexin-45 in mouse retina but not in the developing heart.

The gap junction protein connexin-45 (Cx45) is expressed in the conduction system of the heart and in certain neurons of the retina and brain. General and cardiomyocyte-directed deficiencies of Cx45 in mice lead to lethality on embryonic day 10.5 as a result of cardiovascular defects. Neuron-directed deletion of Cx45 leads to defects in transmission of visual signals. Connexin-36 (Cx36) is co-expressed with Cx45 in certain types of retinal interneurons. To determine whether these two connexins have similar functions and whether Cx36 can compensate for Cx45, we generated knock-in mice in which DNA encoding Cx45 was replaced with that encoding Cx36. Neuron-directed replacement of Cx45 with Cx36 resulted in viable animals. Electroretinographic and neurotransmitter coupling analyses demonstrated functional compensation in the retina. By contrast, general and cardiomyocyte-directed gene replacement led to lethality on embryonic day 11.5. Mutant embryos displayed defects in cardiac morphogenesis and conduction. Thus, functional compensation of Cx45 by Cx36 did not occur during embryonic heart development. These data suggest that Cx45 and Cx36 have similar functions in the retina, whereas Cx45 fulfills special functions in the developing heart that cannot be compensated by Cx36.