Endothelin-Induced Sarcoplasmic Reticulum Calcium Depletion Waves in Vascular Smooth Muscle Cells

Agonist-stimulated waves of elevated cytoplasmic Ca2+ concentration ([Ca2+]i ) regulate blood vessel tone and vasomotion in vascular smooth muscle. Previous studies employing cytoplasmic Ca2+ indicators revealed that these Ca2+ waves were generated by a combination of inositol 1,4,5-trisphosphate (IP3) and Ca2+ induced Ca2+ release (CICR) from the sarcoplasmic reticulum (SR); although, some of the mechanistic details remain uncertain. However, these findings were derived indirectly from observing agonist-induced [Ca2+]i fluctuations in the cytoplasm.
Here, for the first time, we have recorded Endothelin-1 (ET-1) induced waves of Ca2+ depletion from the SR lumen in vascular smooth muscle cells (VSMCs) using a calsequestrin-targeted Ca2+ indicator. Our findings show that these waves: (1) are due to regenerative CICR by the receptors for IP3 (IP3R), (2) have a marked latency period, (3) are characterized by a transient increase in SR Ca2+ ([Ca2+]SR ) both at the point of origin and at the wave front, (4) proceed with diminishing velocity, and (5) are arrested by the nuclear envelope. Our quantitative model indicates that the gradual decrease in the velocity of the SR depletion wave, in the absence of external Ca2+, results from continuity of the SR luminal network

Lorentz-Lorenz Coefficient, Critical Point Constants, and Coexistence Curve of 1,1-Difluoroethylene

We report measurements of the Lorentz-Lorenz coefficient density dependence, the critical temperature, and the critical density, of the fluid 1,1-difluoroethylene. Lorentz-Lorenz coefficient data were obtained by measuring refractive index and density of the same fluid sample independently of one another. Accurate determination of the Lorentz-Lorenz coefficient is necessary for transformation of refractive index data into density data from optics-based experiments on critical phenomena of fluid systems done with different apparatus, with which independent measurement of the refractive indes and density is not possible. Measurements were made along the coexistence curve of the fluid and span the density range 0.01 to 0.80 g/cc. The Lorentz-Lorenz coefficient results show a stronger density dependence along the coexistence curve than previously observed in other fluids, with a monotonic decrease from a density of about 0.2 g/cc onwards, and an overall variation of about 2.5% in the density range studied. No anomaly in the Lorentz-Lorenz coefficient was observed near the critical density. The critical temperature is measured at Tc=(302.964+-0.002) K (29.814 C) and the measured critical density is (0.4195+-0.0018)g/cc.Comment: 14 pages, 6 figures, MikTeX 2.4, submitted to Physical Review

Optical Studies of the coexistence curve of the n-heptane+nitrobenzene mixture near its consolute point measured by an optical method

We have measured the coexistence curve of the binary liquid mixture n-heptane+nitrobenzene near its consolute point using an optical method. In particular, the critical exponent beta describing the coexistence curve was measured for this system. Previous experimental values of beta for n-heptane+nitrobenzene were higher than the typical theoretically calculated value, an unusual, although not unique, occurrence. In an effort to study this discrepancy, we have used an improved experimental apparatus for our measurements. We have taken special care to minimize temperature gradients and maximize the temperature stability of our thermal control system. We have also exploited features of a known optical method to analyze, thoroughly, sources of systematic errors. We measured an apparent value of beta as 0.367+/- 0.006 and by a careful study of the known sources of error we find that they are not able to remove the discrepancy between the measured and the theoretical values of beta. We also measured the critical temperature of the system at Tc=291.80+/- 0.02 K (18.65 C).Comment: 21 pages, 8 figures, submitted to Physical Review B. Shrunk Experimental section, expanded Equilibration and Conclusion sections, eliminated a figure, added reference; 19 pages, 7 figures, resubmitted to PRB. Replaced Fig. 3, added separate simple text file with coexistence curve data (OPvsT.txt); resubmitted to PR

Optical studies of critical phenomena in fluids

Accurate optical techniques were employed to study the behaviour of the binary liquid mixture n-heptane+nitrobenzene (C₇H₁₆ + C₆H₅NO₂) and of the pure fluid 1,1-difluoroethylene (C₂H₂F₂) near their respective critical regions. It is found that, in the region of reduced temperature, t = (T[sub c] — T)/T[sub c] < 3 x 10 ⁻⁴ , the order parameter ΔΦ[sub H] for the binary mixture follows a power law ΔΦ[sub H]« t[sup β], with a leading exponent β = 0.367 ± 0.006, which is higher than the predicted theoretical value of β = 0.326 ± 0.002. A careful study of the effect of refractive index gradients caused by the Earth's gravitational field, potential variation of the optical thickness of the sample cells due to wetting of the cell walls by one the phases, and the long thermal equilibration times of binary mixtures has failed to resolve the disagreement between β measured on n-heptane+nitrobenzene and its theoretical value. A significant feature of this investigation is the novel application of an optical technique (the image plane technique) for studying critical phenomena in transparent binary liquids. The Lorentz-Lorenz function, £, of 1,1-difluoroethylene was measured in order to determine the fluid's density from its refractive index. The quantity £. is found to vary by about 1.4% with density of the fluid, p, with a gentle maximum at a density slightly higher than the critical density, p[sub c]. The density measurements were then used to construct the coexistence curve for this material. The coexistence curve of C₂H₂F₂ was measured with the 'Lorentz-Lorenz' experiment and with a new apparatus combining two complementary optical methods, the prism cell technique and the focal plane technique, into one. The measurements carried out on C₂H₂F₂ in this thesis serve as a test of this apparatus. The order parameter data of C₂H₂F₂ are described accurately by a scaling power law in terms of the reduced temperature t, with the critical exponents at their theoretical values. The coexistence curve data^ taken simultaneously with the two combined techniques agree well with each over the whole range of temperature investigated.Science, Faculty ofPhysics and Astronomy, Department ofGraduat

Two-Dimensional Interfacial Exchange Diffusion Has the Potential to Augment Spatiotemporal Precision of Ca²⁺ Signaling

Nano-junctions between the endoplasmic reticulum and cytoplasmic surfaces of the plasma membrane and other organelles shape the spatiotemporal features of biological Ca²⁺ signals. Herein, we propose that 2D Ca²⁺ exchange diffusion on the negatively charged phospholipid surface lining nano-junctions participates in guiding Ca²⁺ from its source (channel or carrier) to its target (transport protein or enzyme). Evidence provided by in vitro Ca²⁺ flux experiments using an artificial phospholipid membrane is presented in support of the above proposed concept, and results from stochastic simulations of Ca²⁺ trajectories within nano-junctions are discussed in order to substantiate its possible requirements. Finally, we analyze recent literature on Ca²⁺ lipid interactions, which suggests that 2D interfacial Ca²⁺ diffusion may represent an important mechanism of signal transduction in biological systems characterized by high phospholipid surface to aqueous volume ratios.Medicine, Faculty ofNon UBCAnesthesiology, Pharmacology and Therapeutics, Department ofReviewedFacultyResearcherOthe