A unified model for two modes of bursting in GnRH neurons

Gonadotropin-releasing hormone (GnRH) neurons exhibit at least two intrinsic modes of action potential burst firing, referred to as parabolic and irregular bursting. Parabolic bursting is characterized by a slow wave in membrane potential that can underlie periodic clusters of action potentials with increased interspike interval at the beginning and at the end of each cluster. Irregular bursting is characterized by clusters of action potentials that are separated by varying durations of interburst intervals and a relatively stable baseline potential. Based on recent studies of isolated ionic currents, a stochastic Hodgkin-Huxley (HH)-like model for the GnRH neuron is developed to reproduce each mode of burst firing with an appropriate set of conductances. Model outcomes for bursting are in agreement with the experimental recordings in terms of interburst interval, interspike interval, active phase duration, and other quantitative properties specific to each mode of bursting. The model also shows similar outcomes in membrane potential to those seen experimentally when tetrodotoxin (TTX) is used to block action potentials during bursting, and when estradiol transitions cells exhibiting slow oscillations to irregular bursting mode in vitro. Based on the parameter values used to reproduce each mode of bursting, the model suggests that GnRH neurons can switch between the two through changes in the maximum conductance of certain ionic currents, notably the slow inward Ca²⁺ current I _s, and the Ca²⁺ -activated K⁺ current I _KCa. Bifurcation analysis of the model shows that both modes of bursting are similar from a dynamical systems perspective despite differences in burst characteristics.     

Mechanisms of membrane potential oscillation in bursting neurons on the snail,Helix pomatia

• 1.1. The mechanism of generation of membrane potential (MP) oscillations was studied in identified bursting neurons from the snail Helix pomatia.
• 2.2. Long-lasting stimulation of an identified peptidergic interneuron produced a persistent bursting activity in a non-active burster.
• 3.3. External application of calcium channel blockers (1 mM Cd²⁺ or 5 mM La²⁺) resulted in a transient increase in the slow-wave amplitude and subsequent prevention of pacemaker activity generation in bursting neurons. Application of these blockers together with endogenous neuropeptide initiating bursting activity generation, increased MP wave amplitude without prevention of bursting activity generation.
• 4.4. Replacement of all NaCl in normal Ringer's solution with isoosmotic CaCl₂, glucose or Tris-HCl produced a reversible block of bursting activity generation. Stationary current-voltage relation (CVR) of bursting neuron membrane has a region of negative resistance (NRR) and does not intersect the potential axis in threshold region for action potential (AP) generation in normal Ringer's solution. In Na-free solution stationary CVR is linear and intersects the potential axis near — 52 mV.
• 5.5. Novel potential- and time-dependent outward (E_rev = − 58 mV) current, I_B, activated by hyperpolarization was found in the bursting neuron membrane. Having achieved a maximal value, this current decayed with a time constant of about 1 sec. Hyperpolarization inactivated maximal conductance, g_B, responsible for I_B, and depolarization abolished inactivation of g_B.
• 6.6. Short-lasting (0.01 sec) hyperpolarization of the bursting neuron membrane by inward current pulse induced the development of prolonged hyperpolarization wave lasting up to 10 sec.
• 7.7. These results suggest that: (a) persistent bursting activity of RPal neuron in the snail Helix pomatia is not endogenous but is due to a constant activation of peptidergic synaptic inputs of these neurons; (b) Ca²⁺ ions do not play a pivotal role in the ionic mechanism of MP oscillations but play a determining role in the process of secretion of a peptide initiating bursting activity by the interneuron presynaptic terminal; (c) depolarizing phase of the MP wave is due to specific properties of stationary CVR and hyperpolarization phase is due to regenerative properties of hyperpolarization-activated outward current I_B. The minimal mathematical version of MP oscillations based on the experimental data is presented.

     

Ghostbursting: A Novel Neuronal Burst Mechanism

Pyramidal cells in the electrosensory lateral line lobe (ELL) of weakly electric fish have been observed to produce high-frequency burst discharge with constant depolarizing current (Turner et al., 1994). We present a two-compartment model of an ELL pyramidal cell that produces burst discharges similar to those seen in experiments. The burst mechanism involves a slowly changing interaction between the somatic and dendritic action potentials. Burst termination occurs when the trajectory of the system is reinjected in phase space near the ghost of a saddle-node bifurcation of fixed points. The burst trajectory reinjection is studied using quasi-static bifurcation theory, that shows a period doubling transition in the fast subsystem as the cause of burst termination. As the applied depolarization is increased, the model exhibits first resting, then tonic firing, and finally chaotic bursting behavior, in contrast with many other burst models. The transition between tonic firing and burst firing is due to a saddle-node bifurcation of limit cycles. Analysis of this bifurcation shows that the route to chaos in these neurons is type I intermittency, and we present experimental analysis of ELL pyramidal cell burst trains that support this model prediction. By varying parameters in a way that changes the positions of both saddle-node bifurcations in parameter space, we produce a wide gallery of burst patterns, which span a significant range of burst time scales.     

Multiple timescale mixed bursting dynamics in a respiratory neuron model

Experimental results in rodent medullary slices containing the pre-Bötzinger complex (pre-BötC) have identified multiple bursting mechanisms based on persistent sodium current (I _NaP) and intracellular Ca²⁺. The classic two-timescale approach to the analysis of pre-BötC bursting treats the inactivation of I _NaP, the calcium concentration, as well as the Ca²⁺-dependent inactivation of IP ₃ as slow variables and considers other evolving quantities as fast variables. Based on its time course, however, it appears that a novel mixed bursting (MB) solution, observed both in recordings and in model pre-BötC neurons, involves at least three timescales. In this work, we consider a single-compartment model of a pre-BötC inspiratory neuron that can exhibit both I _NaP and Ca²⁺ oscillations and has the ability to produce MB solutions. We use methods of dynamical systems theory, such as phase plane analysis, fast-slow decomposition, and bifurcation analysis, to better understand the mechanisms underlying the MB solution pattern. Rather surprisingly, we discover that a third timescale is not actually required to generate mixed bursting solutions. Through our analysis of timescales, we also elucidate how the pre-BötC neuron model can be tuned to improve the robustness of the MB solution.     

EEG activation response under different neurohumoral states

In order to determine under which neurohumoral conditions the response to usual opening of the eyes stimulates the Berger effect, the electroencephalographic, electrocardiographic, and electromyographic responses to eyes opening have been recorded simultaneously with the psychometric indices of emotional tension and cognitive performance in 59 healthy women aged 18–27 years every two or three days during one or two menstrual cycles determined according the progesterone level in the morning. For excluding the influence of the Novelty factor, the monitoring began at the menstrual phase of the cycle in 29 women and at the luteal phase in the other 30 women. A single examination has been performed in a separate group of 30 women to study the relationship of these parameters with the current progesterone and cortisol levels in saliva. Two-factor ANOVA has shown that the magnitude of amplitude suppression and the bandwidth of the low-frequency α EEG range in the follicular phase of the menstrual cycle are greater than in the luteal one and depend on the Novelty factor. The indices of the Berger effect of the upper-frequency α range do not depend on the neurohumoral state or the Novelty factor. The amplitude suppression and the bandwidth of the lowfrequency α range alone are predictors of the activation in the response to eyes opening because the changes are unidirectional and are interrelated with the autonomic and hormonal characteristics of the activation. It has been demonstrated that eyes opening is a stimulus for the activation only in the neurohumoral state corresponding to the follicular phase of the menstrual cycle. This study has established the dependence of the central and autonomic activation on the individual α frequency EEG profile and the neurohumoral state.     

Modeling the effects of extracellular potassium on bursting properties in pre-Bötzinger complex neurons

There are many types of neurons that intrinsically generate rhythmic bursting activity, even when isolated, and these neurons underlie several specific motor behaviors. Rhythmic neurons that drive the inspiratory phase of respiration are located in the medullary pre-Bötzinger Complex (pre-BötC). However, it is not known if their rhythmic bursting is the result of intrinsic mechanisms or synaptic interactions. In many cases, for bursting to occur, the excitability of these neurons needs to be elevated. This excitation is provided in vitro (e.g. in slices), by increasing extracellular potassium concentration (K _out ) well beyond physiologic levels. Elevated K _out shifts the reversal potentials for all potassium currents including the potassium component of leakage to higher values. However, how an increase in K _out , and the resultant changes in potassium currents, induce bursting activity, have yet to be established. Moreover, it is not known if the endogenous bursting induced in vitro is representative of neural behavior in vivo. Our modeling study examines the interplay between K _out , excitability, and selected currents, as they relate to endogenous rhythmic bursting. Starting with a Hodgkin-Huxley formalization of a pre-BötC neuron, a potassium ion component was incorporated into the leakage current, and model behaviors were investigated at varying concentrations of K _out . Our simulations show that endogenous bursting activity, evoked in vitro by elevation of K _out , is the result of a specific relationship between the leakage and voltage-dependent, delayed rectifier potassium currents, which may not be observed at physiological levels of extracellular potassium.     

Dissection of a model for neuronal parabolic bursting

We have obtained new insight into the mechanisms for bursting in a class of theoretical models. We study Plant's model [24] for Aplysia R-15 to illustrate our view of these so-called parabolic bursters, which are characterized by low spike frequency at the beginning and end of a burst. By identifying and analyzing the fast and slow processes we show how they interact mutually to generate spike activity and the slow wave which underlies the burst pattern. Our treatment is essentially the first step of a singular perturbation approach presented from a geometrical viewpoint and carried out numerically with AUTO [12]. We determine the solution sets (steady state and oscillatory) of the fast subsystem with the slow variables treated as parameters. These solutions form the slow manifold over which the slow dynamics then define a burst trajectory. During the silent phase of a burst, the solution trajectory lies approximately on the steady state branch of the slow manifold and during the active phase of spiking, the trajectory sweeps through the oscillation branch. The parabolic nature of bursting arises from the (degenerate) homoclinic transition between the oscillatory branch and the steady state branch. We show that, for some parameter values, the trajectory remains strictly on the steady state branch (to produce a resting steady state or a pure slow wave without spike activity) or strictly in the oscillatory branch (continuous spike activity without silent phases). Plant's model has two slow variables: a calcium conductance and the intracellular free calcium concentration, which activates a potassium conductance. We also show how bursting arises from an alternative mechanism in which calcium inactivates the calcium current and the potassium conductance is insensitive to calcium. These and other biophysical interpretations are discussed.     

Circadian chronotypic death in heat-synchronized infradian mode cultures ofTetrahymena pyriformis W.

Continuous cultures ofTetrahymena pyriformis in the infradian (“slow-exponential”) growth mode can be entrained to give circadian rhythms of cell division by the application of thermal cycles with periods of 24 hr. The degree of synchronization (phasing index) is dependent on the cycling of other environmental agents, particularly food in the form of the defined medium used. The phase angles of these multiple zeitgebers to one another and to the circadian cell cycle markedly influence the quality of synchronization achieved. In wellsynchronized oxygen-limited cultures, a circadian cytotoxia (circadian chronotypic death) is observed when the culture's oxygen requirements periodically exceed the oxygen support limit of the environment.     

Tracing dynamic biological processes during phase transition

Background

Phase transition widely exists in the biological world, such as transformation of cell cycle phases, cell differentiation stages, disease development, and so on. Such a nonlinear phenomenon is considered as the conversion of a biological system from one phenotype/state to another. Studies on the molecular mechanisms of biological phase transition have attracted much attention, in particular, on different genotypes (or expression variations) in a specific phase, but with less of focus on cascade changes of genes' functions (or system state) during the phase shift or transition process. However, it is a fundamental but important mission to trace the temporal characteristics of a biological system during a specific phase transition process, which can offer clues for understanding dynamic behaviors of living organisms.

Results

By overcoming the hurdles of traditional time segmentation and temporal biclustering methods, a causal process model (CPM) in the present work is proposed to study the biological phase transition in a systematic manner, i.e. first, we make gene-specific segmentation on time-course expression data by developing a new boundary gene estimation scheme, and then infer functional cascade dynamics by constructing a temporal block network. After the computational validation on synthetic data, CPM was used to analyze the well-known Yeast cell cycle data. It was found that the dynamics of the boundary genes are periodic and consistent with the phases of the cell cycle, and the temporal block network indeed demonstrates a meaningful cascade structure of the enriched biological functions. In addition, we further studied protein modules based on the temporal block network, which reflect temporal features in different cycles.

Conclusions

All of these results demonstrate that CPM is effective and efficient comparing to traditional methods, and is able to elucidate essential regulatory mechanism of a biological system even with complicated nonlinear phase transitions.

     

Behavior of a single neuron in a recurrent excitatory loop

A recurrent excitation loop was constructed by enabling each impulse from the slowly adapting stretch receptor organ SAO (crayfish) to trigger through an electronic circuit a brief stretch, or “tug,” of the receptor. When applied independently, each tug influenced the discharge as would an EPSP. Recurrent excitation led to characteristic discharge timings; hence, even an isolated neuron can have intrinsic mechanisms that prevent positive feedback from freezing it in an extreme non-operational state. Such timings depended critically on the “phase”, i.e., on the time elapsed between an SAO impulse and the tug. When the control discharge was stationary (because the SAO length remained invariant), phases of a few ms simply changed the pattern to one of doublets, and affected little the average rate. As the phase increased, bursts appeared, bursts and interburst intervals became more prolonged, and average rates increased. With the largest phases examined (40 ms), the discharge consisted of a slow alternation of high rate bursts, separated by long intervals. When the discharge was modulated (by 0.2/s sinusoidal length variation) with recurrent excitation, the peak-to-peak rate swing, i.e., the sensitivity, and the proportion of the cycle without afferent discharges increased, and the rate vs. length display was distorted even though remaining “loop-plus-extension.” Changes were phase-dependent: for example, loops could have a sharp high peak at one phase and be flat-topped at another. When the interspike interval variability was exaggerated (by a length jitter superimposed upon either invariant or sinusoidally varying lengths), recurrent excitation exerted fewer, weaker and somewhat different effects: e.g., it reduced the overall intensity of the invariant cases and the peak-to-peak swing in the modulated one. The precise mechanisms of these results can only be conjectured at but are likely to involve an electrogenic pump, electromechanical interactions, topographical issues, as well as their interplays. The functional implications involve, for instance, the modulation of the intensity, duration and occurrence of the bursting patterns in oscillating functions (e.g., breathing, chewing, etc.).     

Convergent neuromodulation onto a network neuron can have divergent effects at the network level

Different neuromodulators often target the same ion channel. When such modulators act on different neuron types, this convergent action can enable a rhythmic network to produce distinct outputs. Less clear are the functional consequences when two neuromodulators influence the same ion channel in the same neuron. We examine the consequences of this seeming redundancy using a mathematical model of the crab gastric mill (chewing) network. This network is activated in vitro by the projection neuron MCN1, which elicits a half-center bursting oscillation between the reciprocally-inhibitory neurons LG and Int1. We focus on two neuropeptides which modulate this network, including a MCN1 neurotransmitter and the hormone crustacean cardioactive peptide (CCAP). Both activate the same voltage-gated current (I _MI ) in the LG neuron. However, I _MI-MCN1 , resulting from MCN1 released neuropeptide, has phasic dynamics in its maximal conductance due to LG presynaptic inhibition of MCN1, while I _MI-CCAP retains the same maximal conductance in both phases of the gastric mill rhythm. Separation of time scales allows us to produce a 2D model from which phase plane analysis shows that, as in the biological system, I _MI-MCN1 and I _MI-CCAP primarily influence the durations of opposing phases of this rhythm. Furthermore, I _MI-MCN1 influences the rhythmic output in a manner similar to the Int1-to-LG synapse, whereas I _MI-CCAP has an influence similar to the LG-to-Int1 synapse. These results show that distinct neuromodulators which target the same voltage-gated ion channel in the same network neuron can nevertheless produce distinct effects at the network level, providing divergent neuromodulator actions on network activity.     

Auxin levels in single half nodes ofAvena fatua estimated using high performance liquid chromatography with coulometric detection

A high performance liquid chromatography (HPLC) based micro-method for estimation of indole-3-acetic acid (IAA) levels in single half nodes from the flowering stalks ofAvena fatua has been developed; this features a dual electrode coulometric electrochemical detector operating at a detection limit of c. 2 pg. Samples were prepared by solvent partitioning and preliminary fractionation with C¹⁸ Sep-Pak cartridges. Two stages of reversed phase ion pair HPLC were employed; the first was gradient elution with fluorescent detection, the second, isocratic elution with coulometric detection. The lower limit for estimation of IAA levels in purified extracts was c. 5 pg.     

Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining

Ex vivo ELISPOT and multimer staining are well-established tests for the assessment of antigen-specific T cells. Many laboratories are now using a period of in vitro stimulation (IVS) to enhance detection. Here, we report the findings of a multi-centre panel organised by the Association for Cancer Immunotherapy Immunoguiding Program to investigate the impact of IVS protocols on the detection of antigen-specific T cells of varying ex vivo frequency. Five centres performed ELISPOT and multimer staining on centrally prepared PBMCs from 3 donors, both ex vivo and following IVS. A harmonised IVS protocol was designed based on the best-performing protocol(s), which was then evaluated in a second phase on 2 donors by 6 centres. All centres were able to reliably detect antigen-specific T cells of high/intermediate frequency both ex vivo (Phase I) and post-IVS (Phase I and II). The highest frequencies of antigen-specific T cells ex vivo were mirrored in the frequencies following IVS and in the detection rates. However, antigen-specific T cells of a low/undetectable frequency ex vivo were not reproducibly detected post-IVS. Harmonisation of the IVS protocol reduced the inter-laboratory variation observed for ELISPOT and multimer analyses by approximately 20 %. We further demonstrate that results from ELISPOT and multimer staining correlated after (P < 0.0001 and R ² = 0.5113), but not before IVS. In summary, IVS was shown to be a reproducible method that benefitted from method harmonisation.     

Electronic, ductile, phase transition and mechanical properties of Lu-monopnictides under high pressures

The structural, elastic and electronic properties of lutatium-pnictides (LuN, LuP, LuAs, LuSb, and LuBi) were analyzed by using full-potential linearized augmented plane wave within generalized gradient approximation in the stable rock-salt structure (B1 phase) with space group Fm-3m and high-pressure CsCl structure (B2 phase) with space group Pm-3m. Hubbard-U and spin-orbit coupling were included to predict correctly the semiconducting band gap of LuN. Under compression, these materials undergo first-order structural transitions from B1 to B2 phases at 241, 98, 56.82, 25.2 and 32.3 GPa, respectively. The computed elastic properties show that LuBi is ductile by nature. The electronic structure calculations show that LuN is semiconductor at ambient conditions with an indirect band gap of 1.55 eV while other Lu-pnictides are metallic. It was observed that LuN shows metallization at high pressures. The structural properties, viz, equilibrium lattice constant, bulk modulus and its pressure derivative, transition pressure, equation of state, volume collapse, band gap and elastic moduli, show good agreement with available data.

Cannabinoid receptor activation inhibits cell cycle progression by modulating 14-3-3β

Cannabinoids display various pharmacological activities, including tumor regression, anti-inflammatory and neuroprotective effects. To investigate the molecular mechanisms underlying the pharmacological effects of cannabinoids, we used a yeast two-hybrid system to screen a mouse brain cDNA library for proteins interacting with type 1 cannabinoid receptor (CB1R). Using the intracellular loop 3 of CB1R as bait, we identified 14-3-3β as an interacting partner of CB1R and confirmed their interaction using affinity-binding assays. 14-3-3β has been reported to induce a cell cycle delay at the G₂/M phase. We tested the effects of cannabinoids on cell cycle progression in HeLa cells synchronized using a double-thymidine block-and-release protocol and found an increase in the population of G₂/M phase cells. We further found that CB1R activation augmented the interaction of 14-3-3β with Wee1 and Cdc25B, and promoted phosphorylation of Cdc2 at Tyr-15. These results suggest that cannabinoids induce cell cycle delay at the G2/M phase by activating 14-3-3β.     

Coexistence of Tonic Spiking Oscillations in a Leech Neuron Model

The leech neuron model studied here has a remarkable dynamical plasticity. It exhibits a wide range of activities including various types of tonic spiking and bursting. In this study we apply methods of the qualitative theory of dynamical systems and the bifurcation theory to analyze the dynamics of the leech neuron model with emphasis on tonic spiking regimes. We show that the model can demonstrate bi-stability, such that two modes of tonic spiking coexist. Under a certain parameter regime, both tonic spiking modes are represented by the periodic attractors. As a bifurcation parameter is varied, one of the attractors becomes chaotic through a cascade of period-doubling bifurcations, while the other remains periodic. Thus, the system can demonstrate co-existence of a periodic tonic spiking with either periodic or chaotic tonic spiking. Pontryagins averaging technique is used to locate the periodic orbits in the phase space.     

Multiple modes of a conditional neural oscillator

We present a model for a conditional bursting neuron consisting of five conductances: Hodgkin-Huxley type time- and voltage-dependent Na⁺ and K⁺ conductances, a calcium activated voltage-dependent K⁺ conductance, a calcium-inhibited time- and voltage-dependent Ca⁺⁺ conductance, and a leakage Cl(_– conductance. With an initial set of parameters (versionS), the model shows a hyperpolarized steady-state membrane potential at which the neuron is silent. Increasingg _Na and decreasingg _Cl, whereg _i , is the maximal conductance for speciesi, produces bursts of action potentials (BursterN). Alternatively, an increase ing _Ca produces a different bursting state (BursterC). The two bursting states differ in the periods and amplitudes of their bursting pacemaker potentials. They show different steady-stateI–V curves under simulated voltage-clamp conditions; in simulations that mimic a steady-stateI–V curve taken under experimental conditions only BursterN shows a negative slope resistance region. ModelC continues to burst in the presence of TTX, while bursting in ModelN is suppressed in TTX. Hybrid models show a smooth transition between the two states.     

HNCA+, HNCO+, and HNCACB+ experiments: improved performance by simultaneous detection of orthogonal coherence transfer pathways

Three experiments, BEST–TROSY HNCA+, HNCO+ and HNCACB+ are presented for sequential backbone resonance assignment of ¹³C, ¹⁵N labelled proteins. The novelty of these experiments with respect to conventional pulse sequences is the detection of additional orthogonal coherence transfer pathways that results in enhanced sensitivity for sequential correlations without significantly compromising the intensity of intra-residue correlation peaks. In addition, a 2-step phase cycle separates peaks originating from the orthogonal coherence transfer pathways in 2 sub-spectra, thus providing similar information as obtained from performing a pair of sequential and intra-residue correlation experiments.     

Cytometric analysis of growth-regulator-dependent transcription and cell-cycle progression in Petunia protoplast cultures

Acridine orange simultaneously stains DNA and RNA. Using flow cytometry, synthesis of these nucleic acids can be related throughout a culture time-course. This technique has been used with nuclei isolated from Petunia hybrida protoplasts during 48 h of culture. Nuclear RNA content has been evaluated with respect to DNA levels, namely the cell-cycle phase. Nuclear RNA synthesis was not dependent upon exogeneous hormones during the first 18 h of culture, but either auxin (2,4-dichlorophenoxyacetic acid, 2,4-D) or cytokinin (N⁶-benzyladenine) were necessary for entry into the S phase. Cytokinin alone could stimulate maximal RNA synthesis within each cell-cycle phase up to 24 h. In complete medium, DNA synthesis only began from a phase “G₁B” having substantial RNA, although a subnormal amount of RNA (in protoplasts cultivated only with 2,4-D) did not prevent protoplast entry into the S phase. However, both hormones were necessary for highest RNA levels and G₂ frequencies after 48 h. As in mammalian cells, the mean RNA level in plant 4C nuclei is double that of 2C nuclei. G₂ nuclei are larger than G₁ nuclei, but upon activation G₁ nuclei in fact diminsh in size. This study aimed to identify restriction points in the cell cycle as affected by growth regulators and the specific synthesis of nucleic acids. For example, the RNA levels induced by N⁶-benzyladenine, although similar to those in complete medium, were not sufficient to induce mitosis. Conversely, 2,4-D action was probably limited by low nucleotide synthesis in the absence of cytokinin.