Shaping Neuronal Network Activity by Presynaptic Mechanisms

Neuronal microcircuits generate oscillatory activity, which has been linked to basic functions such as sleep, learning and sensorimotor gating. Although synaptic release processes are well known for their ability to shape the interaction between neurons in microcircuits, most computational models do not simulate the synaptic transmission process directly and hence cannot explain how changes in synaptic parameters alter neuronal network activity. In this paper, we present a novel neuronal network model that incorporates presynaptic release mechanisms, such as vesicle pool dynamics and calcium-dependent release probability, to model the spontaneous activity of neuronal networks. The model, which is based on modified leaky integrate-and-fire neurons, generates spontaneous network activity patterns, which are similar to experimental data and robust under changes in the model''s primary gain parameters such as excitatory postsynaptic potential and connectivity ratio. Furthermore, it reliably recreates experimental findings and provides mechanistic explanations for data obtained from microelectrode array recordings, such as network burst termination and the effects of pharmacological and genetic manipulations. The model demonstrates how elevated asynchronous release, but not spontaneous release, synchronizes neuronal network activity and reveals that asynchronous release enhances utilization of the recycling vesicle pool to induce the network effect. The model further predicts a positive correlation between vesicle priming at the single-neuron level and burst frequency at the network level; this prediction is supported by experimental findings. Thus, the model is utilized to reveal how synaptic release processes at the neuronal level govern activity patterns and synchronization at the network level.     

The activation of the atypical PKC zeta in light‐induced retinal degeneration and its involvement in L‐DNase II control

Light‐induced retinal degeneration is characterized by photoreceptor cell death. Many studies showed that photoreceptor demise is caspase‐independent. In our laboratory we showed that leucocyte elastase inhibitor/LEI‐derived DNase II (LEI/L‐DNase II), a caspase‐independent apoptotic pathway, is responsible for photoreceptor death. In this work, we investigated the activation of a pro‐survival kinase, the protein kinase C (PKC) zeta. We show that light exposure induced PKC zeta activation. PKC zeta interacts with LEI/L‐DNase II and controls its DNase activity by impairing its nuclear translocation. These results highlight the role of PKC zeta in retinal physiology and show that this kinase can control caspase‐independent pathways.     

Universal markers for comparative mapping and phylogenetic analysis in the Asteraceae (Compositae)

The development of universal markers that can be assayed across taxa, but which are polymorphic within taxa, can facilitate both comparative map-based studies and phylogenetic analyses. Here we describe the development of such markers for use in the Asteraceae, which includes the crops lettuce, sunflower, and safflower as well as dozens of locally important crop and weed species. Using alignments of a conserved orthologous set (COS) of ESTs from lettuce and sunflower and genomic sequences of Arabidopsis, we designed a suite of primer pairs that are conserved across species, but which are predicted to flank introns. We then tested 192 such primer pairs in 8 species from across the family. Of these, 163 produced an amplicon in at least 1 taxon, and 125 amplified in at least half of the taxa surveyed. Thirty-nine amplified in all 8 species. Comparisons amongst sequences within the lettuce and sunflower EST databases indicate that the vast majority of these loci will be polymorphic. As a direct test of the utility of these markers outside the lettuce and sunflower subfamilies, we sequenced a subset of ten loci from a panel of cultivated safflower individuals. All 10 loci proved to be single-locus, and nine of the 10 loci were polymorphic with an average of 12.8 SNPs per kb. Taken together, these loci will provide an initial backbone for comparative genetic analyses within the Asteraceae. Moreover, our results indicate that these loci are phylogenetically informative, and hence can be used to resolve evolutionary relationships between taxa within the family as well as within species. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Mark A. Chapman and JianCheng Chang have contributed equally to this work.     

Integrating the effects of area, isolation, and habitat heterogeneity on species diversity: a unification of island biogeography and niche theory

We present an analytical model that unifies two of the most influential theories in community ecology, namely, island biogeography and niche theory. Our model captures the main elements of both theories by incorporating the combined effects of area, isolation, stochastic colonization and extinction processes, habitat heterogeneity, and niche partitioning in a unified, demographically based framework. While classical niche theory predicts a positive relationship between species richness and habitat heterogeneity, our unified model demonstrates that area limitation and dispersal limitation (the main elements of island biogeography) may create unimodal and even negative relationships between species richness and habitat heterogeneity. We attribute this finding to the fact that increasing heterogeneity increases the potential number of species that may exist in a given area (as predicted by niche theory) but simultaneously reduces the amount of suitable area available for each species and, thus, increases the likelihood of stochastic extinction. Area limitation, dispersal limitation, and low reproduction rates intensify the latter effect by increasing the likelihood of stochastic extinction. These analytical results demonstrate that the integration of island biogeography and niche theory provides new insights about the mechanisms that regulate the diversity of ecological communities and generates unexpected predictions that could not be attained from any single theory.     

Stochastic emergence of repeating cortical motifs in spontaneous membrane potential fluctuations in vivo

It was recently discovered that subthreshold membrane potential fluctuations of cortical neurons can precisely repeat during spontaneous activity, seconds to minutes apart, both in brain slices and in anesthetized animals. These repeats, also called cortical motifs, were suggested to reflect a replay of sequential neuronal firing patterns. We searched for motifs in spontaneous activity, recorded from the rat barrel cortex and from the cat striate cortex of anesthetized animals, and found numerous repeating patterns of high similarity and repetition rates. To test their significance, various statistics were compared between physiological data and three different types of stochastic surrogate data that preserve dynamical characteristics of the recorded data. We found no evidence for the existence of deterministically generated cortical motifs. Rather, the stochastic properties of cortical motifs suggest that they appear by chance, as a result of the constraints imposed by the coarse dynamics of subthreshold ongoing activity.     

Placental growth factor contributes to micro-vascular abnormalization and blood-retinal barrier breakdown in diabetic retinopathy

Objective

There are controversies regarding the pro-angiogenic activity of placental growth factor (PGF) in diabetic retinopathy (DR). For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1).

Materials and Methods

pVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes. Control eyes received saline or naked plasmid ET. Clinical follow up was carried out over three months using slit lamp examination and fluorescein angiography. After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts. Ocular structures were visualized through histological analysis.

Results

After fifteen days of rPGF-1 over-expression in normal eyes, tortuous and dilated capillaries were observed. At one month, microaneurysms and moderate vascular sprouts were detected in mid retinal periphery in vivo and on retinal flat-mounts. At later stages, retinal pigmented epithelial cells demonstrated morphological abnormalities and junction ruptures. In diabetic retinas, PGF expression rose between 2 and 5 months, and, one month after ET, rPGF-1 over-expression induced glial activation and proliferation.

Conclusion

This is the first demonstration that sustained intraocular PGF production induces vascular and retinal changes similar to those observed in the early stages of diabetic retinopathy. PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease.     

Agonist-induced phosphorylation and desensitization of the P2Y2 nucleotide receptor

Purification of HA-tagged P2Y₂ receptors from transfected human 1321N1 astrocytoma cells yielded a protein with a molecular size determined by SDS-PAGE to be in the range of 57–76 kDa, which is typical of membrane glycoproteins with heterogeneous complex glycosylation. The protein phosphatase inhibitor, okadaic acid, attenuated the recovery of receptor activity from the agonist-induced desensitized state, suggesting a role for P2Y₂ receptor phosphorylation in desensitization. Isolation of HA-tagged P2Y₂ nucleotide receptors from metabolically [³²P]-labelled cells indicated a (3.8 ± 0.2)-fold increase in the [³²P]-content of the receptor after 15 min of treatment with 100 μM UTP, as compared to immunoprecipitated receptors from untreated control cells. Receptor sequestration studies indicated that ∼40% of the surface receptors were internalized after a 15-min stimulation with 100 μM UTP. Point mutation of three potential GRK and PKC phosphorylation sites in the third intracellular loop and C-terminal tail of the P2Y₂ receptor (namely, S243A, T344A, and S356A) extinguished agonist-induced receptor phosphorylation, caused a marked reduction in the efficacy of UTP to desensitize P2Y₂ receptor signalling to intracellular calcium mobilization, and impaired agonist-induced receptor internalization. Activation of PKC isoforms with phorbol 12-myristate 13-acetate that caused heterologous receptor desensitization did not increase the level of P2Y₂ receptor phosphorylation. Our results indicate a role for receptor phosphorylation by phorbol-insensitive protein kinases in agonist-induced desensitization of the P2Y₂ nucleotide receptor. (Mol Cell Biochem xxx: 35–45, 2005)