Denis Noble

Excitation-Contraction Coupling and Extracellular Calcium Transients in Rabbit Atrium: Reconstruction of Basic Cellular Mechanisms

Interactions of electrogenic sodium-calcium exchange, calcium channel and sarcoplasmic reticulum in the mammalian heart have been explored by simulation of extracellular calcium transients measured with tetramethylmurexide in rabbit atrium. The approach has been to use the simplest possible formulations of these mechanisms, which together with a minimum number of additional mechanisms allow reconstruction of action potentials, intracellular calcium transients and extracellular calcium transients. A 3:1 sodium-calcium exchange stoichiometry is assumed. Calcium-channel inactivation is assumed to take place by a voltage-dependent mechanism, which is accelerated by a rise in intracellular calcium; intracellular calcium release becomes a major physiological regulator of calcium influx via calcium channels. A calcium release mechanism is assumed, which is both calcium- and voltage-sensitive, and which undergoes prolonged inactivation. 200 μm cytosolic calcium buffer is assumed. For most simulations only instantaneous potassium conductances are simulated so as to study the other mechanisms independently of time- and calcium-dependent outward current. Thus, the model reconstructs extracellular calcium transients and typical action-potential configuration changes during steady-state and non-steady-state stimulation from the mechanisms directly involved in trans-sarcolemmal calcium movements. The model predicts relatively small trans-sarcolemmal calcium movements during regular stimulation (ca. 2 μmol kg-1 fresh mass per excitation); calcium current is fully activated within 2 ms of excitation, inactivation is substantially complete within 30 ms, and sodium-calcium exchange significantly resists repolarization from approximately -30 mV. Net calcium movements many times larger are possible during non-steady-state stimulation. Long action potentials at premature excitations or after inhibition of calcium release can be supported almost exclusively by calcium current (net calcium influx 5-30 μmol kg-1 fresh mass); action potentials during potentiated post stimulatory contractions can be supported almost exclusively by sodium-calcium exchange (net calcium efflux 4-20 μmol kg-1 fresh mass). Large calcium movements between the extracellular space and the sarcoplasmic reticulum can take place through the cytosol with virtually no contractile activation. The simulations provide integrated explanations of electrical activity, contractile function and trans-sarcolemmal calcium movements, which were outside the explanatory range of previous models.

Computational physiology and the physiome project

Bioengineering analyses of physiological systems use the computational solution of physical conservation laws on anatomically detailed geometric models to understand the physiological function of intact organs in terms of the properties and behaviour of the cells and tissues within the organ. By linking behaviour in a quantitative, mathematically defined sense across multiple scales of biological organization – from proteins to cells, tissues, organs and organ systems – these methods have the potential to link patient‐specific knowledge at the two ends of these spatial scales. A genetic profile linked to cardiac ion channel mutations, for example, can be interpreted in relation to body surface ECG measurements via a mathematical model of the heart and torso, which includes the spatial distribution of cardiac ion channels throughout the myocardium and the individual kinetics for each of the approximately 50 types of ion channel, exchanger or pump known to be present in the heart. Similarly, linking molecular defects such as mutations of chloride ion channels in lung epithelial cells to the integrated function of the intact lung requires models that include the detailed anatomy of the lungs, the physics of air flow, blood flow and gas exchange, together with the large deformation mechanics of breathing. Organizing this large body of knowledge into a coherent framework for modelling requires the development of ontologies, markup languages for encoding models, and web‐accessible distributed databases. In this article we review the state of the field at all the relevant levels, and the tools that are being developed to tackle such complexity. Integrative physiology is central to the interpretation of genomic and proteomic data, and is becoming a highly quantitative, computer‐intensive discipline.

Simulation of multiple ion channel block provides improved early prediction of compounds' clinical torsadogenic risk

Aims The level of inhibition of the human Ether-à-go-go-related gene (hERG) channel is one of the earliest preclinical markers used to predict the risk of a compound causing Torsade-de-Pointes (TdP) arrhythmias. While avoiding the use of drugs with maximum therapeutic concentrations within 30-fold of their hERG inhibitory concentration 50% (IC50) values has been suggested, there are drugs that are exceptions to this rule: hERG inhibitors that do not cause TdP, and drugs that can cause TdP but are not strong hERG inhibitors. In this study, we investigate whether a simulated evaluation of multi-channel effects could be used to improve this early prediction of TdP risk. Methods and results We collected multiple ion channel data (hERG, Na, l-type Ca) on 31 drugs associated with varied risks of TdP. To integrate the information on multi-channel block, we have performed simulations with a variety of mathematical models of cardiac cells (for rabbit, dog, and human ventricular myocyte models). Drug action is modelled using IC50 values, and therapeutic drug concentrations to calculate the proportion of blocked channels and the channel conductances are modified accordingly. Various pacing protocols are simulated, and classification analysis is performed to evaluate the predictive power of the models for TdP risk. We find that simulation of action potential duration prolongation, at therapeutic concentrations, provides improved prediction of the TdP risk associated with a compound, above that provided by existing markers. Conclusion The suggested calculations improve the reliability of early cardiac safety assessments, beyond those based solely on a hERG block effect.

The IUPS human physiome project

Abstract. The Physiome Project of the International Union of Physiological Sciences (IUPS) is attempting to provide a comprehensive framework for modelling the human body using computational methods which can incorporate the biochemistry, biophysics and anatomy of cells, tissues and organs. A major goal of the project is to use computational modelling to analyse integrative biological function in terms of underlying structure and molecular mechanisms. To support that goal the project is establishing web-accessible physiological databases dealing with model-related data, including bibliographic information, at the cell, tissue, organ and organ system levels. Here we discuss the background and goals of the project, the problems of modelling across multiple spatial and temporal scales, and the development of model ontologies and markup languages at all levels of biological function.

Systems Biology: An Approach

In just over a decade, Systems Biology has moved from being an idea, or rather a disparate set of ideas, to a mainstream feature of research and funding priorities. Institutes, departments, and centers of various flavors of Systems Biology have sprung up all over the world. An Internet search now produces more than 2 million hits. Of the 2,800 entries in PubMed with “Systems Biology” in either the title or the abstract, only two papers were published before 2000, and >90% were published in the past five years. In this article, we interpret Systems Biology as an approach rather than as a field or a destination of research. We illustrate that this approach is productive for the exploration of systems behavior, or “phenotypes,” at all levels of structural and functional complexity, explicitly including the supracellular domain, and suggest how this may be related conceptually to genomes and biochemical networks. We discuss the role of models in Systems Biology and conclude with a consideration of their utility in biomedical research and development.

Requirement of neuronal‐ and cardiac‐type sodium channels for murine sinoatrial node pacemaking

The majority of Na+ channels in the heart are composed of the tetrodotoxin (TTX)‐resistant (KD, 2–6 μm) Nav1.5 isoform; however, recently it has been shown that TTX‐sensitive (KD, 1–10 nm) neuronal Na+ channel isoforms (Nav1.1, Nav1.3 and Nav1.6) are also present and functionally important in the myocytes of the ventricles and the sinoatrial (SA) node. In the present study, in mouse SA node pacemaker cells, we investigated Na+ currents under physiological conditions and the expression of cardiac and neuronal Na+ channel isoforms. We identified two distinct Na+ current components, TTX resistant and TTX sensitive. At 37°C, TTX‐resistant iNa and TTX‐sensitive iNa started to activate at ∼−70 and ∼−60 mV, and peaked at −30 and −10 mV, with a current density of 22 ± 3 and 18 ± 1 pA pF−1, respectively. TTX‐sensitive iNa inactivated at more positive potentials as compared to TTX‐resistant iNa. Using action potential clamp, TTX‐sensitive iNa was observed to activate late during the pacemaker potential. Using immunocytochemistry and confocal microscopy, different distributions of the TTX‐resistant cardiac isoform, Nav1.5, and the TTX‐sensitive neuronal isoform, Nav1.1, were observed: Nav1.5 was absent from the centre of the SA node, but present in the periphery of the SA node, whereas Nav1.1 was present throughout the SA node. Nanomolar concentrations (10 or 100 nm) of TTX, which block TTX‐sensitive iNa, slowed pacemaking in both intact SA node preparations and isolated SA node cells without a significant effect on SA node conduction. In contrast, micromolar concentrations (1–30 μm) of TTX, which block TTX‐resistant iNa as well as TTX‐sensitive iNa, slowed both pacemaking and SA node conduction. It is concluded that two Na+ channel isoforms are important for the functioning of the SA node: neuronal (putative Nav1.1) and cardiac Nav1.5 isoforms are involved in pacemaking, although the cardiac Nav1.5 isoform alone is involved in the propagation of the action potential from the SA node to the surrounding atrial muscle.

The rise of computational biology

The year 2001 saw a remarkable burst of interest in biological simulation, with several international meetings on the subject, and the inclusion, by journals, of web site references from which published models can be downloaded. So, why has all this happened so suddenly?

Claude Bernard, the first systems biologist, and the future of physiology

The first systems analysis of the functioning of an organism was Claude Bernards concept of the constancy of the internal environment (le milieu intérieur), since it implied the existence of control processes to achieve this. He can be regarded, therefore, as the first systems biologist. The new vogue for systems biology today is an important development, since it is time to complement reductionist molecular biology by integrative approaches. Claude Bernard foresaw that this would require the application of mathematics to biology. This aspect of Claude Bernards work has been neglected by physiologists, which is why we are not as ready to contribute to the development of systems biology as we should be. In this paper, I outline some general principles that could form the basis of systems biology as a truly multilevel approach from a physiologists standpoint. We need the insights obtained from higher‐level analysis in order to succeed even at the lower levels. The reason is that higher levels in biological systems impose boundary conditions on the lower levels. Without understanding those conditions and their effects, we will be seriously restricted in understanding the logic of living systems. The principles outlined are illustrated with examples from various aspects of physiology and biochemistry. Applying and developing these principles should form a major part of the future of physiology.

A theory of biological relativity: no privileged level of causation

Must higher level biological processes always be derivable from lower level data and mechanisms, as assumed by the idea that an organism is completely defined by its genome? Or are higher level properties necessarily also causes of lower level behaviour, involving actions and interactions both ways? This article uses modelling of the heart, and its experimental basis, to show that downward causation is necessary and that this form of causation can be represented as the influences of initial and boundary conditions on the solutions of the differential equations used to represent the lower level processes. These insights are then generalized. A priori, there is no privileged level of causation. The relations between this form of ‘biological relativity’ and forms of relativity in physics are discussed. Biological relativity can be seen as an extension of the relativity principle by avoiding the assumption that there is a privileged scale at which biological functions are determined.

Sodium‐calcium exchange during the action potential in guinea‐pig ventricular cells.

1. Slow inward tail currents attributable to electrogenic sodium‐calcium exchange can be recorded by imposing hyperpolarizing voltage clamp pulses during the normal action potential of isolated guinea‐pig ventricular cells. The hyperpolarizations return the membrane to the resting potential (between ‐65 and ‐88 m V) allowing an inward current to be recorded. This current usually has peak amplitude when repolarization is imposed during the first 50 ms after the action potential upstroke, but becomes negligible once the final phase of repolarization is reached. The envelope of peak current tail amplitudes strongly resembles that of the intracellular calcium transient recorded in other studies. 2. Repetitive stimulation producing normal action potentials at a frequency of 2 Hz progressively augments the tail current recorded immediately after the stimulus train. Conversely, if each action potential is prematurely terminated at 0.1 Hz, repetitive stimulation produces a tail current much smaller than the control value. The control amplitude of inward current is only maintained if interrupted action potentials are separated by at least one full ‘repriming’ action potential. These effects mimic those on cell contraction (Arlock & Wohlfart, 1986) and suggest that progressive changes in tail current are controlled by variations in the amplitude and time course of the intracellular calcium transient. 3. When intracellular calcium is buffered sufficiently to abolish contraction, the tail current is abolished. Substitution of calcium with strontium greatly reduces the tail current. 4. The inward tail current can also be recorded at more positive membrane potentials using standard voltage clamp pulse protocols. In this way it was found that temperature has a large effect on the tail current, which can change from net inward at 22 degrees C to net outward at 37 degrees C. The largest inward currents are usually recorded at about 30 degrees C. It is shown that this effect is attributable predominantly to the temperature sensitivity of activation of the delayed potassium current, iK, whose decay can then mask the slow tail current at high temperatures. 5. Studies of the relationship between the tail current and the membrane calcium current, iCa, have been performed using a method of drug application which is capable of perturbing iCa in a very rapid and highly reversible manner. Partial block of iCa with cadmium does not initially alter the size of the associated inward current tail. When iCa is increased by applying isoprenaline, the percentage augmentation of the associated tail current is much greater but occurs more slowly.(ABSTRACT TRUNCATED AT 400 WORDS)