MAP65/Ase1 promote microtubule flexibility.

Microtubules (MTs) are dynamic cytoskeletal elements involved in numerous cellular processes. Although they are highly rigid polymers with a persistence length of 1-8 mm, they may exhibit a curved shape at a scale of few micrometers within cells, depending on their biological functions. However, how MT flexural rigidity in cells is regulated remains poorly understood. Here we ask whether MT-associated proteins (MAPs) could locally control the mechanical properties of MTs. We show that two major cross-linkers of the conserved MAP65/PRC1/Ase1 family drastically decrease MT rigidity. Their MT-binding domain mediates this effect. Remarkably, the softening effect of MAP65 observed on single MTs is maintained when MTs are cross-linked. By reconstituting physical collisions between growing MTs/MT bundles, we further show that the decrease in MT stiffness induced by MAP65 proteins is responsible for the sharp bending deformations observed in cells when they coalign at a steep angle to create bundles. Taken together, these data provide new insights into how MAP65, by modifying MT mechanical properties, may regulate the formation of complex MT arrays.

In vivo measurement of lung capillary-alveolar macromolecule permeability by saturation bronchoalveolar lavage.

OBJECTIVE: Measurement of capillary-alveolar permeability to fluorescein isothiocyanate-dextran (FITC-D) (molecular mass, 71,300 daltons) by a sequential bronchoalveolar lavage (BAL) technique. DESIGN: Animal research. SETTING: The Department of Physiology at a scientific and medical university. SUBJECTS: Nine anesthetized and mechanically ventilated dogs. INTERVENTIONS: Two separate experiments were performed in each subject-an initial control experiment followed by an oleic acid-induced lung injury. The indicator was administered at constant blood concentration before serial BAL including eight fluid instillation-recovery cycles. MEASUREMENTS: Plasma to BAL solute clearance at saturation (capillary-alveolar clearance at saturation, mL/min) was calculated and normalized to lavage fluid volume (measured by 1251 serum albumin dilution) to obtain a transport rate (TR) constant. MAIN RESULTS: TR for FITC-D70 was 4.0+/-0.8 and 46.1+/-18.1 x 10(-5) x min(-1) in control and injured lung, respectively (p < .02). Capillary-alveolar clearance of FITC-D70 was not affected by the lavage procedure itself. TR reflected essentially epithelial permeability in normal lung and combined epithelial and endothelial permeability in injured lung. A significant correlation was found between cardiac output and TR in injured lung. CONCLUSIONS: Saturation BAL allowed us to estimate capillary-alveolar macromolecule permeability in vivo in dogs. Further study may allow bedside evaluation of lung injury by BAL in patients.

Implications of relaxation dynamics in the synaptic control of olfactory cortex activity.

In a previous work (Ballain et al., 1998. Biol. Cyber. 79, 323-336) we reported the analysis of a model for the piriform cortex activity in rats based on experimental data. In this paper, we study an extension of this model by supplementing it with equations for the post-synaptic conductance and/or the pre-synaptic activation threshold. We use the present model's outputs to account for experimental data based on paired stimulation in the opossum or the rat, obtained either through electrical recording or optical mapping of the cortex activity. The model exhibits great robustness when it comes to large variation in synaptic characteristics. Model outputs mimic satisfactorily the three kind of responses to paired stimuli (Litaudon and Cattarelli, 1996. Eur. J. Neurosci. 8, 21-29) and the recovery of the excitable capacities as demonstrated by Haberly (1973. J. Neurophysiol. 36 (4), 789-802) and Ferreyra-Moyano et al. (1985. Brain Res. Bull. 15, 237 248).

Role of the net architecture in piriform cortex activity: analysis by a mathematical model.

We present a mathematical analysis of the piriform cortex activity in rats. Experimental data were obtained by means of optical recording of fluorescent signals driven by neuronal activity. From these data, we determined the numerical value of the relaxation time for the pyramidal cell activity in layers II and III and the time latency map for bulb activation. Our model for the piriform cortex is based on pairs of excitatory and inhibitory neurons which correspond to pyramidal cells of layers II and III and to their inhibitory associated interneurons respectively; pyramidal cells are also interconnected through short and long range association fiber systems. Under such conditions, the model outputs resemble closely the experimental observations: (1) a double-bumped response to a strong and short stimulation; (2) oscillatory behavior under weak sustained stimulation conditions; (3) propagation of traveling activity waves; and (4) pacemaker activity when clusters of neurons are preferentially coupled.

Coexistence of multiple propagating wave-fronts in a regulated enzyme reaction model: link with birhythmicity and multi-threshold excitability.

We analyze the spatial propagation of wave-fronts in a biochemical model for a product-activated enzyme reaction with non-linear recycling of product into substrate. This model was previously studied as a prototype for the coexistence of two distinct types of periodic oscillations (birhythmicity). The system is initially in a stable steady state characterized by the property of multi-threshold excitability, by which it is capable of amplifying in a pulsatory manner perturbations exceeding two distinct thresholds. In such conditions, when the effect of diffusion is taken into account, two distinct wave-fronts are shown to propagate in space, with distinct amplitudes and velocities, for the same set of parameter values, depending on the magnitude of the initial perturbation. Such a multiplicity of propagating wave-fronts represents a new type of coexistence of multiple modes of dynamic behavior, besides the coexistence involving, under spatially homogeneous conditions, multiple steady states, multiple periodic regimes, or a combination of steady and periodic regimes.

Modelling nonlinear integration of synaptic signals by neurones.

Neurones with active conductance on dendrites integrate synaptic signals and modulate generation of axon spikes in a nonlinear way. Owing to experimental difficulties, modelling provides invaluable insight for the comprehension of neurone behaviour particularly when dendrites are excitable. We used experimental data obtained for the Anterior Gastric Receptor neurone (AGR neurone), which controls the lobster gastric mill activity, to derive a set of partial differential equations for the membrane voltage. Simulation showed that upon varying the intensity of stimulation on the dendrite, the response pattern between dendrites and axon activity continuously changes. In addition, when only half of the dendritic tree is active, axon firing exhibits regular oscillations and bursting activity. We discuss these results in relation with the experimental work done on the AGR neurone.

Modeling the integrative properties of dendrites: application to the striatal spiny neuron.

The role of the striatum in the control of movements and in the processing of cortical information has received much attention in the recent years. We set out a simple biophysical model for the medium-spiny neuron (msn), the most abundant cell in striatum. This neuron receives two main kinds of inputs, namely, cortical excitatory inputs and dopaminergic inputs coming from the substantia nigra pars compacta. The msn axon impinges directly onto the globus pallidus and onto the substantia nigra pars reticulata neurons and onto striatal neurons through recurrent branches of the axon. The msn is characterized by spiny dendritic trees with a high density of spines (1 to 4 spines/microns) and the probable existence of dendritic spikes. The model predicts that the neuron can integrate excitable inputs in a linear or a nonlinear mode. In the nonlinear mode, the neuron allows the detection of simultaneous (or almost simultaneous) synaptic inputs; it facilitates either a slowing down or a speeding up of the information transfer between the synaptic input location and the soma and is sensitive to inhibiton-excitation pairing. Conversely, in the linear integrative mode, the somatic voltage is determined by a weighted summation of the synaptic inputs. Several geometrical, electrical, or temporal factors can control the switch between these behaviors: the density of excitable dendritic elements, the dendritic radius, the resistance of the spine stem, the membrane resistance, the time between excitations, and the distance between synaptic sites. Finally, the signification of this behavior is discussed in connection with the putative role of dopamine and with the striatal net organization.

Kinetics of endosomal pH evolution in Dictyostelium discoideum amoebae. Study by fluorescence spectroscopy.

The evolution of endo-lysosomal pH in Dictyostelium discoideum amoebae was examined during fluid-phase endocytosis. Pulse-chase experiments were conducted in nutritive medium or in non-nutritive medium using fluorescein labelled dextran (FITC-dextran) as fluid-phase marker and pH probe. In both conditions, efflux kinetics were characterized by an extended lag phase lasting for 45-60 min and corresponding to intracellular transit of FITC-dextran cohort. During the chase period, endosomal pH decreased during approximately 20 min from extracellular pH down to pH 4.6-5.0, then, it increased within the next 20-40 min to reach pH 6.0-6.2. It was only at this stage that FITC-dextran was released back into the medium with pseudo first-order kinetics. A vacuolar H(+)-ATPase is involved in endosomal acidification as the acidification process was markedly reduced in mutant strain HGR8, partially defective in vacuolar H(+)-ATPase and in parent type strain AX2 by bafilomycin A1, a selective inhibitor of this enzyme. Our data suggest that endocytic cargo is channeled from endosomes to secondary lysosomes that are actively linked to the plasma membrane via recycling vesicles.

Suppression of chaos and other dynamical transitions induced by intercellular coupling in a model for cyclic AMP signaling in Dictyostelium cells.

The effect of intercellular coupling on the switching between periodic behavior and chaos is investigated in a model for cAMP oscillations in Dictyostelium cells. We first analyze the dynamic behavior of a homogeneous cell population which is governed by a three-variable differential system for which bifurcation diagrams are obtained as a function of two control parameters. We then consider the mixing of two populations behaving in a chaotic and periodic manner, respectively. Cells are coupled through the sharing of a common chemical intermediate, extracellular cAMP, which controls its production and release by the cells into the extracellular medium; the dynamics of the mixed suspension is governed by a five-variable differential system. When the two cell populations differ by the value of a single parameter which measures the activity of the enzyme that degrades extracellular cAMP, the bifurcation diagram established for the three-variable homogeneous population can be used to predict the dynamic behavior of the mixed suspension. The analysis shows that a small proportion of periodic cells can suppress chaos in the mixed suspension. Such a fragility of chaos originates from the relative smallness of the domain of aperiodic oscillations in parameter space. The bifurcation diagram is used to obtain the minimum fraction of periodic cells suppressing chaos. These results are related to the suppression of chaos by the small-amplitude periodic forcing of a strange attractor. Numerical simulations further show how the coupling of periodic cells with chaotic cells can produce chaos, bursting, simple periodic oscillations, or a stable steady state; the coupling between two populations at steady state can produce similar modes of dynamic behavior.

Suppression of chaos by periodic oscillations in a model for cyclic AMP signalling in Dictyostelium cells.

We investigate how the introduction of cells oscillating periodically affects the behaviour of a suspension of Dictyostelium discoideum amoebae undergoing chaotic oscillations of cyclic AMP. The analysis of a model indicates that a tiny proportion of periodic cells suffices to transform chaos into periodic oscillations in such suspensions. A similar result is obtained by forcing the aperiodic oscillations by a small-amplitude, periodic input of cyclic AMP. The results provide an explanation for the observation of regular oscillations in suspensions of a putatively chaotic mutant of Dictyostelium discoideum. More generally, the results show how chaos in biological systems may disappear through the coupling with periodic oscillations.

Finding complex oscillatory phenomena in biochemical systems. An empirical approach.

Starting with a model for a product-activated enzymatic reaction proposed for glycolytic oscillations, we show how more complex oscillatory phenomena may develop when the basic model is modified by addition of product recycling into substrate or by coupling in parallel or in series two autocatalytic enzyme reactions. Among the new modes of behavior are the coexistence between two stable types of oscillations (birhythmicity), bursting, and aperiodic oscillations (chaos). On the basis of these results, we outline an empirical method for finding complex oscillatory phenomena in autonomous biochemical systems, not subjected to forcing by a periodic input. This procedure relies on finding in parameter space two domains of instability of the steady state and bringing them close to each other until they merge. Complex phenomena occur in or near the region where the two domains overlap. The method applies to the search for birhythmicity, bursting and chaos in a model for the cAMP signalling system of Dictyostelium discoideum amoebae.

A Model Based on Receptor Desensitization for Cyclic AMP Signaling in Dictyostelium Cells.

We analyze a model based on receptor modification for the cAMP signaling system that controls aggregation of the slime mold Dictyostelium discoideum after starvation. The model takes into account both the desensitization of the cAMP receptor by reversible phosphorylation and the activation of adenylate cyclase that follows binding of extracellular cAMP to the unmodified receptor. The dynamics of the signaling system is studied in terms of three variables, namely, intracellular and extracellular cAMP, and the fraction of receptor in active state. Using parameter values collected from experimental studies on cAMP signaling and receptor phosphorylation, we show that the model accounts qualitatively and, in a large measure, quantitatively for the various modes of dynamic behavior observed in the experiments: (a) autonomous oscillations of cAMP, (b) relay of suprathreshold cAMP pulses, i.e., excitability, characterized by both an absolute and a relative refractory period, and (c) adaptation to constant cAMP stimuli. A two-variable version of the model is used to demonstrate the link between excitability and oscillations by phase plane analysis. The response of the model to repetitive stimulation allows comprehension, in terms of receptor desensitization, of the role of periodic signaling in Dictyostelium and, more generally, the function of pulsatile patterns of hormone secretion.

Autonomous chaotic behaviour of the slime mould Dictyostelium discoideum predicted by a model for cyclic AMP signalling.

How sustained oscillations lose their periodicity and thus give rise to chaos was first analysed in mathematical models, then observed in chemical systems such as the Belousov-Zhabotinsky reaction where chaos is autonomous because it originates from endogenous kinetic mechanisms. In contrast, chaos can also be obtained by periodically forcing an oscillatory system, as shown, for example, in cardiac cells and yeast glycolysis. Biochemical evidence for autonomous chaos has been obtained both in vitro for the peroxidase reaction and in enzymatic models not based directly on experimental systems. We report here the occurrence of autonomous chaos in a realistic model for the cyclic AMP signalling system of the slime mould Dictyostelium discoideum, based on receptor modification. This model is also capable of bursting, a phenomenon characteristic of some pacemaker neurones such as R15 in Aplysia. Whereas bursting has not been observed in D. discoideum, our model suggests that 'aperiodic signalling' in the mutant Fr17 provides the first example of autonomous chaos occurring spontaneously at the cellular level.

Metabolic oscillations in biochemical systems controlled by covalent enzyme modification.

We analyze the conditions under which sustained oscillations develop in a biochemical system regulated autocatalytically by reversible, covalent enzyme modification. The analysis applies, for example, to the situation where adenylate cyclase (or guanylate cyclase) is activated through phosphorylation by a cAMP (or cGMP)-dependent protein kinase. The model then provides a non-allosteric mechanism for the periodic generation of cAMP or cGMP pulses. For certain parameter values close to those that produce oscillations, the system is excitable since it can amplify in a pulsatory manner suprathreshold perturbations. The results on excitable and oscillatory behavior are discussed in relation with the mechanism of cAMP relay and oscillation in the slime mold Dictyostelium discoideum.