Enhanced virtual microscopy for collaborative education

<p>Abstract</p> <p>Background</p> <p>Curricular reform efforts and a desire to use novel educational strategies that foster student collaboration are challenging the traditional microscope-based teaching of histology. Computer-based histology teaching tools and Virtual Microscopes (VM), computer-based digital slide viewers, have been shown to be effective and efficient educational strategies. We developed an open-source VM system based on the Google Maps engine to transform our histology education and introduce new teaching methods. This VM allows students and faculty to collaboratively create content, annotate slides with markers, and it is enhanced with social networking features to give the community of learners more control over the system.</p> <p>Results</p> <p>We currently have 1,037 slides in our VM system comprised of 39,386,941 individual JPEG files that take up 349 gigabytes of server storage space. Of those slides 682 are for general teaching and available to our students and the public; the remaining 355 slides are used for practical exams and have restricted access. The system has seen extensive use with 289,352 unique slide views to date. Students viewed an average of 56.3 slides per month during the histology course and accessed the system at all hours of the day. Of the 621 annotations added to 126 slides 26.2% were added by faculty and 73.8% by students. The use of the VM system reduced the amount of time faculty spent administering the course by 210 hours, but did not reduce the number of laboratory sessions or the number of required faculty. Laboratory sessions were reduced from three hours to two hours each due to the efficiencies in the workflow of the VM system.</p> <p>Conclusions</p> <p>Our virtual microscope system has been an effective solution to the challenges facing traditional histopathology laboratories and the novel needs of our revised curriculum. The web-based system allowed us to empower learners to have greater control over their content, as well as the ability to work together in collaborative groups. The VM system saved faculty time and there was no significant difference in student performance on an identical practical exam before and after its adoption. We have made the source code of our VM freely available and encourage use of the publically available slides on our website.</p

A Mathematical model for Astrocytes mediated LTP at Single Hippocampal Synapses

Many contemporary studies have shown that astrocytes play a significant role in modulating both short and long form of synaptic plasticity. There are very few experimental models which elucidate the role of astrocyte over Long-term Potentiation (LTP). Recently, Perea & Araque (2007) demonstrated a role of astrocytes in induction of LTP at single hippocampal synapses. They suggested a purely pre-synaptic basis for induction of this N-methyl-D- Aspartate (NMDA) Receptor-independent LTP. Also, the mechanisms underlying this pre-synaptic induction were not investigated. Here, in this article, we propose a mathematical model for astrocyte modulated LTP which successfully emulates the experimental findings of Perea & Araque (2007). Our study suggests the role of retrograde messengers, possibly Nitric Oxide (NO), for this pre-synaptically modulated LTP.Comment: 51 pages, 15 figures, Journal of Computational Neuroscience (to appear

Calmodulin Activation by Calcium Transients in the Postsynaptic Density of Dendritic Spines

The entry of calcium into dendritic spines can trigger a sequence of biochemical reactions that begins with the activation of calmodulin (CaM) and ends with long-term changes to synaptic strengths. The degree of activation of CaM can depend on highly local elevations in the concentration of calcium and the duration of transient increases in calcium concentration. Accurate measurement of these local changes in calcium is difficult because the spaces are so small and the numbers of molecules are so low. We have therefore developed a Monte Carlo model of intracellular calcium dynamics within the spine that included calcium binding proteins, calcium transporters and ion channels activated by voltage and glutamate binding. The model reproduced optical recordings using calcium indicator dyes and showed that without the dye the free intracellular calcium concentration transient was much higher than predicted from the fluorescent signal. Excitatory postsynaptic potentials induced large, long-lasting calcium gradients across the postsynaptic density, which activated CaM. When glutamate was released at the synapse 10 ms before an action potential occurred, simulating activity patterns that strengthen hippocampal synapses, the calcium gradient and activation of CaM in the postsynaptic density were much greater than when the order was reversed, a condition that decreases synaptic strengths, suggesting a possible mechanism underlying the induction of long-term changes in synaptic strength. The spatial and temporal mechanisms for selectivity in CaM activation demonstrated here could be used in other signaling pathways

Spatial Distribution of Calcium-Gated Chloride Channels in Olfactory Cilia

Background: In vertebrate olfactory receptor neurons, sensory cilia transduce odor stimuli into changes in neuronal membrane potential. The voltage changes are primarily caused by the sequential openings of two types of channel: a cyclic-nucleotide-gated (CNG) cationic channel and a calcium-gated chloride channel. In frog, the cilia are 25 to 200 mm in length, so the spatial distributions of the channels may be an important determinant of odor sensitivity. Principal Findings: To determine the spatial distribution of the chloride channels, we recorded from single cilia as calcium was allowed to diffuse down the length of the cilium and activate the channels. A computational model of this experiment allowed an estimate of the spatial distribution of the chloride channels. On average, the channels were concentrated in a narrow band centered at a distance of 29 % of the ciliary length, measured from the base of the cilium. This matches the location of the CNG channels determined previously. This non-uniform distribution of transduction proteins is consistent with similar findings in other cilia. Conclusions: On average, the two types of olfactory transduction channel are concentrated in the same region of the cilium

Autoinhibition of TBCB regulates EB1-mediated microtubule dynamics

Tubulin cofactors (TBCs) participate in the folding, dimerization, and dissociation pathways of the tubulin dimer. Among them, TBCB and TBCE are two CAP-Gly domain-containing proteins that interact and dissociate the tubulin dimer. Here we show how TBCB localizes at spindle and midzone microtubules during mitosis. Furthermore, the motif DEI/M-COO– present in TBCB, which is similar to the EEY/F-COO– element characteristic of EB proteins, CLIP-170, and α-tubulin, is required for TBCE–TBCB heterodimer formation and thus for tubulin dimer dissociation. This motif is responsible for TBCB autoinhibition, and our analysis suggests that TBCB is a monomer in solution. Mutants of TBCB lacking this motif are derepressed and induce microtubule depolymerization through an interaction with EB1 associated to microtubule tips. TBCB is also able to bind to the chaperonin complex CCT containing α-tubulin, suggesting that it could escort tubulin to facilitate its folding and dimerization, recycling or degradation

IFT Proteins Accumulate during Cell Division and Localize to the Cleavage Furrow in Chlamydomonas

Intraflagellar transport (IFT) proteins are well established as conserved mediators of flagellum/cilium assembly and disassembly. However, data has begun to accumulate in support of IFT protein involvement in other processes elsewhere in the cell. Here, we used synchronous cultures of Chlamydomonas to investigate the temporal patterns of accumulation and localization of IFT proteins during the cell cycle. Their mRNAs showed periodic expression that peaked during S and M phase (S/M). Unlike most proteins that are synthesized continuously during G1 phase, IFT27 and IFT46 levels were found to increase only during S/M phase. During cell division, IFT27, IFT46, IFT72, and IFT139 re-localized from the flagella and basal bodies to the cleavage furrow. IFT27 was further shown to be associated with membrane vesicles in this region. This localization pattern suggests a role for IFT in cell division

Social Motility in African Trypanosomes

African trypanosomes are devastating human and animal pathogens that cause significant human mortality and limit economic development in sub-Saharan Africa. Studies of trypanosome biology generally consider these protozoan parasites as individual cells in suspension cultures or in animal models of infection. Here we report that the procyclic form of the African trypanosome Trypanosoma brucei engages in social behavior when cultivated on semisolid agarose surfaces. This behavior is characterized by trypanosomes assembling into multicellular communities that engage in polarized migrations across the agarose surface and cooperate to divert their movements in response to external signals. These cooperative movements are flagellum-mediated, since they do not occur in trypanin knockdown parasites that lack normal flagellum motility. We term this behavior social motility based on features shared with social motility and other types of surface-induced social behavior in bacteria. Social motility represents a novel and unexpected aspect of trypanosome biology and offers new paradigms for considering host-parasite interactions

The Golgin GMAP210/TRIP11 Anchors IFT20 to the Golgi Complex

Eukaryotic cells often use proteins localized to the ciliary membrane to monitor the extracellular environment. The mechanism by which proteins are sorted, specifically to this subdomain of the plasma membrane, is almost completely unknown. Previously, we showed that the IFT20 subunit of the intraflagellar transport particle is localized to the Golgi complex, in addition to the cilium and centrosome, and hypothesized that the Golgi pool of IFT20 plays a role in sorting proteins to the ciliary membrane. Here, we show that IFT20 is anchored to the Golgi complex by the golgin protein GMAP210/Trip11. Mice lacking GMAP210 die at birth with a pleiotropic phenotype that includes growth restriction, ventricular septal defects of the heart, omphalocele, and lung hypoplasia. Cells lacking GMAP210 have normal Golgi structure, but IFT20 is no longer localized to this organelle. GMAP210 is not absolutely required for ciliary assembly, but cilia on GMAP210 mutant cells are shorter than normal and have reduced amounts of the membrane protein polycystin-2 localized to them. This work suggests that GMAP210 and IFT20 function together at the Golgi in the sorting or transport of proteins destined for the ciliary membrane

Lobe Specific Ca2+-Calmodulin Nano-Domain in Neuronal Spines: A Single Molecule Level Analysis

Calmodulin (CaM) is a ubiquitous Ca2+ buffer and second messenger that affects cellular function as diverse as cardiac excitability, synaptic plasticity, and gene transcription. In CA1 pyramidal neurons, CaM regulates two opposing Ca2+-dependent processes that underlie memory formation: long-term potentiation (LTP) and long-term depression (LTD). Induction of LTP and LTD require activation of Ca2+-CaM-dependent enzymes: Ca2+/CaM-dependent kinase II (CaMKII) and calcineurin, respectively. Yet, it remains unclear as to how Ca2+ and CaM produce these two opposing effects, LTP and LTD. CaM binds 4 Ca2+ ions: two in its N-terminal lobe and two in its C-terminal lobe. Experimental studies have shown that the N- and C-terminal lobes of CaM have different binding kinetics toward Ca2+ and its downstream targets. This may suggest that each lobe of CaM differentially responds to Ca2+ signal patterns. Here, we use a novel event-driven particle-based Monte Carlo simulation and statistical point pattern analysis to explore the spatial and temporal dynamics of lobe-specific Ca2+-CaM interaction at the single molecule level. We show that the N-lobe of CaM, but not the C-lobe, exhibits a nano-scale domain of activation that is highly sensitive to the location of Ca2+ channels, and to the microscopic injection rate of Ca2+ ions. We also demonstrate that Ca2+ saturation takes place via two different pathways depending on the Ca2+ injection rate, one dominated by the N-terminal lobe, and the other one by the C-terminal lobe. Taken together, these results suggest that the two lobes of CaM function as distinct Ca2+ sensors that can differentially transduce Ca2+ influx to downstream targets. We discuss a possible role of the N-terminal lobe-specific Ca2+-CaM nano-domain in CaMKII activation required for the induction of synaptic plasticity