Analysis of pharmacologically isolated components of the ERG

AbstractAn harmonic analysis was applied to the electroretinogram (ERG) measured in intact cat eyes in control conditions and after pharmacological isolation of the components attributed to photoreceptors (PIII) and bipolar neurons (PII). The frequency response curves obtained in various conditions showed that the bandwidth of the PII component extends over a range of stimulus frequencies higher than the bandwidth of PIII. The enhancement of the PII response to stimuli of high temporal frequency suggests the presence of a frequency dependent gain control located either pre- and/or post-synaptically in the transmission line between the phototransductive cascade and bipolar neurons. A possible role of these processes is to enhance relevant visual information whilst selectively attenuating low frequency signals originating in the transductive cascade

Involvement of Autophagic Pathway in the Progression of Retinal Degeneration in a Mouse Model of Diabetes

The notion that diabetic retinopathy (DR) is essentially a micro-vascular disease has been recently challenged by studies reporting that vascular changes are preceded by signs of damage and loss of retinal neurons. As to the mode by which neuronal death occurs, the evidence that apoptosis is the main cause of neuronal loss is far from compelling. The objective of this study was to investigate these controversies in a mouse model of streptozotocin (STZ) induced diabetes. Starting from 8 weeks after diabetes induction there was loss of rod but not of cone photoreceptors, together with reduced thickness of the outer and inner synaptic layers. Correspondingly, rhodopsin expression was downregulated and the scotopic electroretinogram (ERG) is suppressed. In contrast, cone opsin expression and photopic ERG response were not affected. Suppression of the scotopic ERG preceded morphological changes as well as any detectable sign of vascular alteration. Only sparse apoptotic figures were detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and glia was not activated. The physiological autophagy flow was altered instead, as seen by increased LC3 immunostaining at the level of outer plexiform layer (OPL) and upregulation of the autophagic proteins Beclin-1 and Atg5. Collectively, our results show that the streptozotocin induced DR in mouse initiates with a functional loss of the rod visual pathway. The pathogenic pathways leading to cell death develop with the initial dysregulation of autophagy well before the appearance of signs of vascular damage and without strong involvement of apoptosis

Modulating brain integrative actions as a new perspective on pharmacological approaches to neuropsychiatric diseases

: A critical aspect of drug development in the therapy of neuropsychiatric diseases is the "Target Problem", that is, the selection of a proper target after not simply the etiopathological classification but rather the detection of the supposed structural and/or functional alterations in the brain networks. There are novel ways of approaching the development of drugs capable of overcoming or at least reducing the deficits without triggering deleterious side effects. For this purpose, a model of brain network organization is needed, and the main aspects of its integrative actions must also be established. Thus, to this aim we here propose an updated model of the brain as a hyper-network in which i) the penta-partite synapses are suggested as key nodes of the brain hyper-network and ii) interacting cell surface receptors appear as both decoders of signals arriving to the network and targets of central nervous system diseases. The integrative actions of the brain networks follow the "Russian Doll organization" including the micro (i.e., synaptic) and nano (i.e., molecular) levels. In this scenario, integrative actions result primarily from protein-protein interactions. Importantly, the macromolecular complexes arising from these interactions often have novel structural binding sites of allosteric nature. Taking G protein-coupled receptors (GPCRs) as potential targets, GPCRs heteromers offer a way to increase the selectivity of pharmacological treatments if proper allosteric drugs are designed. This assumption is founded on the possible selectivity of allosteric interventions on G protein-coupled receptors especially when organized as "Receptor Mosaics" at penta-partite synapse level

Exosomes from astrocyte processes: Signaling to neurons

open13It is widely recognized that extracellular vesicles subserve non-classical signal transmission in the central nervous system. Here we assess if the astrocyte processes, that are recognized to play crucial roles in intercellular communication at the synapses and in neuron-astrocyte networks, could convey messages through extracellular vesicles. Our findings indicate, for the first time that freshly isolated astrocyte processes prepared from adult rat cerebral cortex, can indeed participate to signal transmission in central nervous system by releasing exosomes that by volume transmission might target near or longdistance sites. It is noteworthy that the exosomes released from the astrocyte processes proved ability to selectively target neurons. The astrocyte-derived exosomes were proven positive for neuroglobin, a protein functioning as neuroprotectant against cell insult; the possibility that exosomes might transfer neuroglobin to neurons would add a mechanism to the potential astrocytic neuroprotectant activity. Notably, the exosomes released from the processes of astrocytes maintained markers, which prove their parental astrocytic origin. This potentially allows the assessment of the cellular origin of exosomes that might be recovered from body fluids.openVenturini A.; Passalacqua M.; Pelassa S.; Pastorino F.; Tedesco M.; Cortese K.; Gagliani M.C.; Leo G.; Maura G.; Guidolin D.; Agnati L.F.; Marcoli M.; Cervetto C.Venturini, A.; Passalacqua, M.; Pelassa, S.; Pastorino, F.; Tedesco, M.; Cortese, K.; Gagliani, M. C.; Leo, G.; Maura, G.; Guidolin, D.; Agnati, L. F.; Marcoli, M.; Cervetto, C

Mechanisms of photoreceptor death and survival in mammalian retina

The mammalian retina, like the rest of the central nervous system, is highly stable and can maintain its structure and function for the full life of the individual, in humans for many decades. Photoreceptor dystrophies are instances of retinal instability. Many are precipitated by genetic mutations and scores of photoreceptor-lethal mutations have now been identified at the codon level. This review explores the factors which make the photoreceptor more vulnerable to small mutations of its proteins than any other cell of the body, and more vulnerable to environmental factors than any other retinal neurone. These factors include the highly specialised structure and function of the photoreceptors, their high appetite for energy, their self-protective mechanisms and the architecture of their energy supply from the choroidal circulation. Particularly important are the properties of the choroidal circulation, especially its fast flow of near-arterial blood and its inability to autoregulate. Mechanisms which make the retina stable and unstable are then reviewed in three different models of retinal degeneration, retinal detachment, photoreceptor dystrophy and light damage. A two stage model of the genesis of photoreceptor dystrophies is proposed, comprising an initial "depletion" stage caused by genetic or environmental insult and a second "late" stage during which oxygen toxicity damages and eventually destroys any photoreceptors which survive the initial depletion. It is a feature of the model that the second "late" stage of retinal dystrophies is driven by oxygen toxicity. The implications of these ideas for therapy of retinal dystrophies are discussed

Processing of Retinal Signals in Normal and HCN Deficient Mice

This study investigates the role of two different HCN channel isoforms in the light response of the outer retina. Taking advantage of HCN-deficient mice models and of in vitro (patch-clamp) and in vivo (ERG) recordings of retinal activity we show that HCN1 and HCN2 channels are expressed at distinct retinal sites and serve different functions. Specifically, HCN1 operate mainly at the level of the photoreceptor inner segment from where, together with other voltage sensitive channels, they control the time course of the response to bright light. Conversely, HCN2 channels are mainly expressed on the dendrites of bipolar cells and affect the response to dim lights. Single cell recordings in HCN1−/− mice or during a pharmacological blockade of Ih show that, contrary to previous reports, Ikx alone is able to generate the fast initial transient in the rod bright flash response. Here we demonstrate that the relative contribution of Ih and Ikx to the rods' temporal tuning depends on the membrane potential. This is the first instance in which the light response of normal and HCN1- or HCN2-deficient mice is analyzed in single cells in retinal slice preparations and in integrated full field ERG responses from intact animals. This comparison reveals a high degree of correlation between single cell current clamp data and ERG measurements. A novel picture emerges showing that the temporal profile of the visual response to dim and bright luminance changes is separately determined by the coordinated gating of distinct voltage dependent conductances in photoreceptors and bipolar cells

High-Pass Filtering of Input Signals by the Ih Current in a Non-Spiking Neuron, the Retinal Rod Bipolar Cell

Hyperpolarization–activated cyclic nucleotide–sensitive (HCN) channels mediate the If current in heart and Ih throughout the nervous system. In spiking neurons Ih participates primarily in different forms of rhythmic activity. Little is known, however, about its role in neurons operating with graded potentials as in the retina, where all four channel isoforms are expressed. Intriguing evidence for an involvement of Ih in early visual processing are the side effects reported, in dim light or darkness, by cardiac patients treated with HCN inhibitors. Moreover, electroretinographic recordings indicate that these drugs affect temporal processing in the outer retina. Here we analyzed the functional role of HCN channels in rod bipolar cells (RBCs) of the mouse. Perforated–patch recordings in the dark–adapted slice found that RBCs exhibit Ih, and that this is sensitive to the specific blocker ZD7288. RBC input impedance, explored by sinusoidal frequency–modulated current stimuli (0.1–30 Hz), displays band–pass behavior in the range of Ih activation. Theoretical modeling and pharmacological blockade demonstrate that high–pass filtering of input signals by Ih, in combination with low–pass filtering by passive properties, fully accounts for this frequency–tuning. Correcting for the depolarization introduced by shunting through the pipette–membrane seal, leads to predict that in darkness Ih is tonically active in RBCs and quickens their responses to dim light stimuli. Immunohistochemistry targeting candidate subunit isoforms HCN1–2, in combination with markers of RBCs (PKC) and rod–RBC synaptic contacts (bassoon, mGluR6, Kv1.3), suggests that RBCs express HCN2 on the tip of their dendrites. The functional properties conferred by Ih onto RBCs may contribute to shape the retina's light response and explain the visual side effects of HCN inhibitors