Dynamics and stimulus-dependence of pacemaker control during behavioral modulations in the weakly electric fish,Apteronotus

1. Weakly electric fish generate around their bodies low-amplitude, AC electric fields which are used both for the detection of objects and intraspecific communication. The types of modulation in this signal of which the high-frequency wave-type gymnotiform, Apteronotus, is capable are relatively few and stereotyped. Chief among these is the chirp, a signal used in courtship and agonistic displays. Chirps are brief and rapid accelerations in the normally highly regular electric organ discharge (EOD) frequency. 2. Chirping can be elicited artificially in these animals by the use of a stimulus regime identical to that typically used to elicit another behavior, the jamming avoidance response (JAR). The neuronal basis for the JAR, a much slower and lesser alteration in EOD frequency, is well understood. Examination of the stimulus features which induce chirping show that, like the JAR, there is a region of frequency differences between the fish's EOD and the interfering signal that maximally elicits the response. Moreover, the response is sex-specific with regard to the sign of the frequency difference, with females chirping preferentially on the positive and most males on the negative Df. These features imply that the sensory mechanisms involved in the triggering of these communicatory behaviors are fundamentally similar to those explicated for the JAR. 3. Additionally, two other modulatory behaviors of unknown significance are described. The first is a non-selective rise in EOD frequency associated with a JAR stimulus, occurring regardless of the sign of the Df. This modulation shares many characteristics with the JAR. The second behavior, which we have termed a 'yodel', is distinct from and kinetically intermediate to chirping and the JAR. Moreover, unlike the other studied electromotor behaviors it is generally produced only after the termination of the eliciting stimulus.     

Russell body phenotype is preferentially induced by IgG mAb clones with high intrinsic condensation propensity: Relations between the biosynthetic events in the ER and solution behaviors in vitro

The underlying reasons for why some mAb (monoclonal antibody) clones are much more inclined to induce a Russell body (RB) phenotype during immunoglobulin biosynthesis remain elusive. Although RBs are morphologically understood as enlarged globular aggregates of immunoglobulins deposited in the endoplasmic reticulum (ER), little is known about the properties of the RB-inducing mAb clones as secretory cargo and their physical behaviors in the extracellular space. To elucidate how RB-inducing propensities, secretion outputs, and the intrinsic physicochemical properties of individual mAb clones are interrelated, we used HEK293 cells to study the biosynthesis of 5 human IgG mAbs for which prominent solution behavior problems were known a priori. All 5 model mAbs with inherently high condensation propensities induced RB phenotypes both at steady state and under ER-to-Golgi transport block, and resulted in low secretion titer. By contrast, one reference mAb that readily crystallized at neutral pH in vitro produced rod-shaped crystalline bodies in the ER without inducing RBs. Another reference mAb without notable solution behavior issues did not induce RBs and was secreted abundantly. Intrinsic physicochemical properties of individual IgG clones thus directly affected the biosynthetic steps in the ER, and thereby produced distinctive cellular phenotypes and influenced IgG secretion output. The findings implicated that RB formation represents a phase separation event or a loss of colloidal stability in the secretory pathway organelles. The process of RB induction allows the cell to preemptively reduce the extracellular concentration of potentially pathogenic, highly aggregation-prone IgG clones by selectively storing them in the ER.     

Russell body phenotype is preferentially induced by IgG mAb clones with high intrinsic condensation propensity: Relations between the biosynthetic events in the ER and solution behaviors in vitro

The underlying reasons for why some mAb (monoclonal antibody) clones are much more inclined to induce a Russell body (RB) phenotype during immunoglobulin biosynthesis remain elusive. Although RBs are morphologically understood as enlarged globular aggregates of immunoglobulins deposited in the endoplasmic reticulum (ER), little is known about the properties of the RB-inducing mAb clones as secretory cargo and their physical behaviors in the extracellular space. To elucidate how RB-inducing propensities, secretion outputs, and the intrinsic physicochemical properties of individual mAb clones are interrelated, we used HEK293 cells to study the biosynthesis of 5 human IgG mAbs for which prominent solution behavior problems were known a priori. All 5 model mAbs with inherently high condensation propensities induced RB phenotypes both at steady state and under ER-to-Golgi transport block, and resulted in low secretion titer. By contrast, one reference mAb that readily crystallized at neutral pH in vitro produced rod-shaped crystalline bodies in the ER without inducing RBs. Another reference mAb without notable solution behavior issues did not induce RBs and was secreted abundantly. Intrinsic physicochemical properties of individual IgG clones thus directly affected the biosynthetic steps in the ER, and thereby produced distinctive cellular phenotypes and influenced IgG secretion output. The findings implicated that RB formation represents a phase separation event or a loss of colloidal stability in the secretory pathway organelles. The process of RB induction allows the cell to preemptively reduce the extracellular concentration of potentially pathogenic, highly aggregation-prone IgG clones by selectively storing them in the ER.     

The reticulon and DP1/Yop1p proteins form immobile oligomers in the tubular endoplasmic reticulum

We recently identified a class of membrane proteins, the reticulons and DP1/Yop1p, which shape the tubular endoplasmic reticulum (ER) in yeast and mammalian cells. These proteins are highly enriched in the tubular portions of the ER and virtually excluded from other regions. To understand how they promote tubule formation, we characterized their behavior in cellular membranes and addressed how their localization in the ER is determined. Using fluorescence recovery after photobleaching, we found that yeast Rtn1p and Yop1p are less mobile in the membrane than normal ER proteins. Sucrose gradient centrifugation and cross-linking analyses show that they form oligomers. Mutants of yeast Rtn1p, which no longer localize exclusively to the tubular ER or are even totally inactive in inducing ER tubules, are more mobile and oligomerize less extensively. The mammalian reticulons and DP1 are also relatively immobile and can form oligomers. The conserved reticulon homology domain that includes the two membrane-embedded segments is sufficient for the localization of the reticulons to the tubular ER, as well as for their diffusional immobility and oligomerization. Finally, ATP depletion in both yeast and mammalian cells further decreases the mobilities of the reticulons and DP1. We propose that oligomerization of the reticulons and DP1/Yop1p is important for both their localization to the tubular domains of the ER and for their ability to form tubules.     

Langevin equation,Fokker-Planck equation and cell migration

Cell migration can be characterized by two independent variables: the speed,v, and the migration angle, ϕ. Each variable can be described by a stochastic differential equation—a Langevin equation. The migration behaviour of an ensemble of cells can be predicted due to the stochastic processes involved in the signal transduction/response system of each cell. Distribution functions, correlation functions, etc. are determined by using the corresponding Fokker-Planck equation. The model assumptions are verified by experimental results. The theoretical predictions are mainly compared with the galvanotactic response of human granulocytes. The coefficient characterizing the mean effect of the signal transduction/response system of the cell is experimentally determined to 0.08 mm/V sec (galvanotaxis) or 0.7 mm/sec (chemotaxis) and the characteristic time characterizing stochastic effects in the signal transduction/response system is experimentally determined as 30 sec. The temporal directed response induced by electric field pulses is investigated: the experimental cells react slower but are more sensitive than predicted by theory.     

Numerical Simulation of Dynamic Electro-Mechanical Response of Ionic Polymer-Metal Composites

Ionic Polymer-Metal Composites (IPMC) is an emerging class of Electro-Active Polymer (EAP) materials. IPMC has attractive features, such as high sensitivity and light weight, which are useful for developing novel designs in the fields of bionic actuators, artificial muscles and dynamic sensors. A Finite Element (FE) model was developed for simulating the dynamic electro-mechanical response of an IPMC structure under an external voltage input. A lumped Resistor-Capacitor (RC) model was used to describe the voltage-to-current relationship of a Nafion IPMC film for the computation of electric field intensity. Moreover, the viscoelastic property of the IPMC film was considered in the model and the non-uniform bending behavior was also taken into account. Based on the proposed model and the assumption that the thicknesses of the two electrodes are the same and uniform, the optimal coating thickness of the IPMC electrode was determined. It was demonstrated that the dynamic electro-mechanical response of the IPMC structure can be predicted by the proposed FE model, and the simulation results were in good agreement with the experimental findings.     

Sensory cues for the gradual frequency fall responses of the gymnotiform electric fish, Rhamphichthys rostratus

The sensory cues for a less known form of frequency shifting behavior, gradual frequency falls, of electric organ discharges (EODs) in a pulse-type gymnotiform electric fish, Rhamphichthys rostratus, were identified. We found that the gradual frequency fall occurs independently of more commonly observed momentary phase shifting behavior, and is due to perturbation of sensory feedback of the fish's own EODs by EODs of neighboring fish. The following components were identified as essential features in the signal mixture of the fish's own and the neighbor's EOD pulses: (1) the neighbor's pulses must be placed within a few millisecond of the fish's own pulses, (2) the neighbor's pulses, presented singly at low frequencies (0.2–4 Hz), were sufficient, (3) the frequency of individual pulse presentation must be below 4 Hz, (4) amplitude modulation of the sensory feedback of the fish's own pulses induced by such insertions of the neighbor's pulses must contain a high frequency component: sinusoidal amplitude modulation of the fish's own EOD feedback at these low frequencies does not induce gradual frequency falls. Differential stimulation across body surfaces, which is required for the jamming avoidance response (JAR) of wave-type gymnotiform electric fish, was not necessary for this behavior. We propose a cascade of high-pass and low-pass frequency filters within the amplitude processing pathway in the central nervous system as the mechanism of the gradual frequency fall response.Abbreviations EOD electric organ discharge - f frequency of EOD or pacemaker command signal - JAR jamming avoidance response - S ₁ stimulus mimicking fish's own EOD - f ₁ frequency of S₁ - S ₂ stimulus mimicking neighbor's EOD - f ₂ frequency of S₂     

Resonant ac-dc magnetic fields: calculated response

An elementary model consisting of one charged particle in a viscous medium exposed to weak ac-dc low-frequency magnetic fields is analyzed to identify and explain the fundamental characteristics of the physical mechanisms that result in a resonance response, which is similar to the familiar cyclotron resonance. The model predicts both frequency and amplitude windows, which are explained in terms of synchronization of the particle with electric fields. Although extrapolation of model results to biological systems is limited by the elementary nature of the model, the model results indicate that observed resonant responses by others of biological systems to ac-dc magnetic fields are probably not due to resonant response of ions in solution, since the model predicts that no resonant response is possible unless the viscous damping is very low, many orders of magnitude lower than the viscous damping of ions in solution.     

An ephaptic transmission model of CA3 pyramidal cells: an investigation into electric field effects

Extracellular electric fields existing throughout the living brain affect the neural coding and information processing via ephaptic transmission, independent of synapses. A two-compartment whole field effect model (WFEM) of pyramidal neurons embedded within a resistive array which simulates the extracellular medium i.e. ephapse is developed to study the effects of electric field on neuronal behaviors. We derive the two linearized filed effect models (LFEM-1 and LFEM-2) from WFEM at the stable resting state. Through matching these simplified models to the subthreshold membrane response in experiments of the resting pyramidal cells exposed to applied electric fields, we not only verify our proposed model’s validity but also found the key parameters which dominate subthreshold frequency response characteristic. Moreover, we find and give its underlying biophysical mechanism that the unsymmetrical properties of active ion channels results in the very different low-frequency response of somatic and dendritic compartments. Following, WFEM is used to investigate both direct-current (DC) and alternating-current field effect on the neural firing patterns by bifurcation analyses. We present that DC electric field could modulate neuronal excitability, with the positive field improving the excitability, the modest negative field suppressing the excitability, but interestingly, the larger negative field re-exciting the neuron back into spiking behavior. The neuron exposed to the sinusoidal electric field exhibits abundant firing patterns sensitive to the input frequency and intensity. In addition, the electrical properties of ephapse can modulate the efficacy of field effect. Our simulated results are qualitatively in line with the relevant experimental results and can explain some experimental phenomena. Furthermore, they are helpful to provide the predictions which can be tested in future experiments.     

Gates and oscillators II: zeitgebers and the network model of the brain clock

Circadian rhythms in physiology and behavior are regulated by the SCN. When assessed by expression of clock genes, at least 2 distinct functional cell types are discernible within the SCN: nonrhythmic, light-inducible, retinorecipient cells and rhythmic autonomous oscillator cells that are not directly retinorecipient. To predict the responses of the circadian system, the authors have proposed a model based on these biological properties. In this model, output of rhythmic oscillator cells regulates the activity of the gate cells. The gate cells provide a daily organizing signal that maintains phase coherence among the oscillator cells. In the absence of external stimuli, this arrangement yields a multicomponent system capable of producing a self-sustained consensus rhythm. This follow-up study considers how the system responds when the gate cells are activated by an external stimulus, simulating a response to an entraining (or phase-setting) signal. In this model, the authors find that the system can be entrained to periods within the circadian range, that the free-running system can be phase shifted by timed activation of the gate, and that the phase response curve for activation is similar to that observed when animals are exposed to a light pulse. Finally, exogenous triggering of the gate over a number of days can organize an arrhythmic system, simulating the light-dependent reappearance of rhythmicity in a population of disorganized, independent oscillators. The model demonstrates that a single mechanism (i.e., the output of gate cells) can account for not only free-running and entrained rhythmicity but also other circadian phenomena, including limits of entrainment, a PRC with both delay and advance zones, and the light-dependent reappearance of rhythmicity in an arrhythmic animal.     

A soluble form of lymphocyte activation gene-3 (IMP321) induces activation of a large range of human effector cytotoxic cells

The principal antitumor immune response is mediated through the activation of type 1 cytotoxic (Tc1) CD8 T cells, NK cells, and monocytes/macrophages. In this study, we investigated the potency of a clinical-grade soluble form of lymphocyte activation gene-3 protein (IMP321), a physiological high-affinity MHC class II binder, at inducing in PBMCs an appropriate cytotoxic-type response in short-term ex vivo assays. We found that IMP321 binds to a minority (<10%) of MHC class II + cells in PBMCs, including all myeloid dendritic cells, and a small fraction of monocytes. Four hours after addition of IMP321 to PBMCs, these myeloid cells produce TNF-alpha and CCL4 as determined by intracellular staining. At 18 h, 1% of CD8+ T cells and 3.7% NK cells produce Tc1 cytokines such as IFN-gamma and/or TNF-alpha (mean values from 60 blood donors). Similar induction was observed in metastatic cancer patient PBMCs, but the values were lower for the NK cell subset. Early APC activation by IMP321 is needed for this Tc1-type activation because pure sorted CD8+ T cells could not be activated by IMP321. Only Ag-experienced, fully differentiated granzyme+ CD8 T cells (effector and effector memory but not naive or central memory T cells) are induced by IMP321 to full Tc1 activation. In contrast to IMP321, TLR1-9 agonists induce IL-10 and are therefore unable to induce this Tc1 IFN-gamma+ response. Thus, IMP321 has many properties that confirm its potential to be a new class of immunopotentiator in cancer patients.     

Cutaneous electrical oscillation in a weakly electric fish, Gymnarchus niloticus

An African electric fish, Gymnarchus niloticus. ceases its electric organ discharge for a prolonged time in response to external electrical signals. During the cessation of electric organ discharges from the electric organ, a weak sinusoidal signal (approximately 0.1 mV cm(-1)) near the fish's previous discharge frequency was recorded near the body. The oscillatory potentials at all points on the body surface were synchronized and had a complex spatial distribution. The source of the potential was determined to be within the dermal tissue. Electroreceptive central neurons that responded to a moving target near the fish with normal electric organ discharges also responded to the same target when the electric organ discharge was interrupted and the potential from the skin existed. This result suggests that the fish may be able to electrolocate objects without the discharge from the electric organ.     

High yield recombinant silk-like protein production in transgenic plants through protein targeting

DP1B is a synthetic analogue of spider dragline silk protein. It can be spun to form silk fiber. Previously, it had been expressed in transgenic plants, showing the general feasibility of the plant-based DP1B production. However, success of such a plant-based platform requires a great increase of DP1B productivity in plant cells to reduce production cost. This report describes a protein targeting approach to accumulate DP1B in apoplast, ER lumen, and vacuole in Arabidopsis cells, by utilizing appropriate combinations of sporamin-targeting determinant peptides and ER retention peptide. The approach has dramatically enhanced DP1B accumulation, resulting in high production yield. The accumulation can be as high as 8.5 and 6.7% total soluble protein in leaf tissue by targeting to apoplast and ER lumen, respectively, or as high as 18 and 8.2% total soluble protein in seeds by targeting to ER lumen and vacuole, respectively. However, the vacuole targeting in leaves and the apoplast targeting in seeds have failed to accumulate full length DP1B molecules or any DP1B at all, respectively, suggesting that they may not be suitable for applications in leaf tissues and seeds. Data in this study recommend a combination of seed-specific expression and ER-targeting as one of the best strategies for yield enhancement of plant-based DP1B production.     

Sine wave electropermeabilization reveals the frequency-dependent response of the biological membranes

The permeabilization of biological membranes by electric fields, known as electroporation, has been traditionally performed with square electric pulses. These signals distribute the energy applied to cells in a wide frequency band. This paper investigates the use of sine waves, which are narrow band signals, to provoke electropermeabilization and the frequency dependence of this phenomenon.Single bursts of sine waves at different frequencies in the range from 8 kHz–130 kHz were applied to cells in vitro. Electroporation was studied in the plasma membrane and the internal organelles membrane using calcium as a permeabilization marker. Additionally, a double-shell electrical model was simulated to give a theoretical framework to our results.The electroporation efficiency shows a low pass filter frequency dependence for both the plasma membrane and the internal organelles membrane. The mismatch between the theoretical response and the observed behavior for the internal organelles membrane is explained by a two-step permeabilization process: first the permeabilization of the external membrane and afterwards that of the internal membranes. The simulations in the model confirm this two-step hypothesis when a variable plasma membrane conductivity is considered in the analysis.This study demonstrates how the use of narrow-band signals as sine waves is a suitable method to perform electroporation in a controlled manner. We suggest that the use of this type of signals could bring a simplification in the investigations of the very complex phenomenon of electroporation, thus representing an interesting option in future fundamental studies.     

Neural adaptation facilitates oscillatory responses to static inputs in a recurrent network of ON and OFF cells

We investigate the role of adaptation in a neural field model, composed of ON and OFF cells, with delayed all-to-all recurrent connections. As external spatially profiled inputs drive the network, ON cells receive inputs directly, while OFF cells receive an inverted image of the original signals. Via global and delayed inhibitory connections, these signals can cause the system to enter states of sustained oscillatory activity. We perform a bifurcation analysis of our model to elucidate how neural adaptation influences the ability of the network to exhibit oscillatory activity. We show that slow adaptation encourages input-induced rhythmic states by decreasing the Andronov–Hopf bifurcation threshold. We further determine how the feedback and adaptation together shape the resonant properties of the ON and OFF cell network and how this affects the response to time-periodic input. By introducing an additional frequency in the system, adaptation alters the resonance frequency by shifting the peaks where the response is maximal. We support these results with numerical experiments of the neural field model. Although developed in the context of the circuitry of the electric sense, these results are applicable to any network of spontaneously firing cells with global inhibitory feedback to themselves, in which a fraction of these cells receive external input directly, while the remaining ones receive an inverted version of this input via feedforward di-synaptic inhibition. Thus the results are relevant beyond the many sensory systems where ON and OFF cells are usually identified, and provide the backbone for understanding dynamical network effects of lateral connections and various forms of ON/OFF responses.     

Model for receptor-controlled cytosolic calcium oscillations and for external influences on the signal pathway.

The external stimulation of many cells by a hormone, for example, often leads to an oscillating cytosolic calcium concentration. This periodic behavior is now designated the cytosolic calcium oscillator. A theoretical model is presented that describes this behavior on the basis of inositol(1,4,5)trisphosphate-induced calcium oscillations. In contrast to other models only a single positive feedback loop is taken into account to obtain oscillations. The model includes important innovations compared to other approaches. It includes the contribution of extracellular calcium and its modification after the stimulation of the cell. Furthermore, the signal pathway that leads to cytosolic calcium oscillations is described in more detail than in other models. This enables investigations on the influence of additional parameters like external electromagnetic fields on the signal transduction pathway. The model and the calculations are based on the theory of nonlinear self-sustained oscillators.     

Retention of a type II surface membrane protein in the endoplasmic reticulum by the Lys-Asp-Glu-Leu sequence.

Soluble luminal proteins of the endoplasmic reticulum (ER) are known to be retained by a tetrapeptide retention signal, KDEL. We report in this communication that the KDEL sequence when appended to the carboxy terminus of a cell surface membrane protein, dipeptidyl peptidase IV (DPPIV), resulted in its retention in the endoplasmic reticulum of transfected Madin-Darby canine kidney cells as assessed by indirect immunofluorescence. Selective surface biotinylation revealed that about 90-95% of the expressed DPPIV was retained in the ER. Appendance of the sequence KDEV did not, however, result in ER retention, illustrating the functional specificity of the retention signal. The ER retention was not due to misfolding of the mutant protein, as the mutant proteins remained enzymatically active. Our data suggest that the KDEL receptor is able to recognize and recycle type II membrane proteins containing a carboxyl-terminal KDEL sequence and postulates the existence of such yet to be identified endogenous proteins.