Hydroxamic Acids

We have studied the effect of ascorbic acid on voltage-dependent calcium channels in pancreatic [3 cells. Using the whole-cell and perforated-patch variants of the patch clamp technique to record calcium tail currents, we have shown that the slowly deactivating (SD) calcium channel, which is similar to the T-type channel in other cells, is inhibited in a voltage-dependent manner by ascorbic acid (AA). The other channels that carry inward current in 13 cells, FD calcium channels and sodium channels, are unaffected by AA. Ascorbic acid causes a voltagedependent decrease in the magnitude of the SD channel conductance which can be explained by the hypothesis that ~ 50-60% of the channels have their voltage dependence shifted by ~ 62 mV in the depolarizing direction. Thus, ascorbate appears to modify only a fraction of the SD channels. The activation kinetics of the ascorbate-modified channels are slower than control channels in a manner that is consistent with this hypothesis. Deactivation and inactivation kinetics are unaffected by ascorbate. These effects of ascorbate require metal ions, and it appears that some of the activity of ascorbate is due to a product of its metal catalyzed oxidation, perhaps dehydroascorbate.

Section: The Ascorbate Effect Requires Heavy Metalmentioning

confidence: 95%

Ascorbic acid modulation of calcium channels in pancreatic beta cells.

Parsey

Matteson

1993

“…Junction potentials were calculated using the Henderson (1907) equation using mobility and activity coefficients from Robinson and Stokes (1965), Weast (1987), andLandolt-B6rnstein (1960). Apparent binding constants for EDTA, EGTA, and ATP were derived from Martell and Smith (1974).…”

Section: Recording Techniquesmentioning

confidence: 99%

Intrinsic gating of inward rectifier in bovine pulmonary artery endothelial cells in the presence or absence of internal Mg2+.

Silver

DeCoursey

1990

120

Inward rectifier (IR) currents were studied in bovine pulmonary artery endothelial cells in the whole-cell configuration of the patch-clamp technique with extracellular K + concentrations, [K+]o, ranging from 4.5 to 160 mM. Whether the concentration of free Mg ~+ in the intraceUular solution, [Mg~+]i , was 1.9 mM or nominally 0, the IR exhibited voltage-and time-dependent gating. The IR conductance was activated by hyperpolarization and deactivated by depolarization. Small steady-state outward IR currents were present up to ~40 mV more positive than the K + reversal potential, EK, regardless of [Mg~+]i. Modeled as a first-order C ~ O gating process, both the opening rate, a, and the closing rate,/3, were exponentially dependent on voltage, with/3 more steeply voltage dependent, changing e-fold for 9 mV compared with 18 mV for an e-fold change in a. Over all [K+]o studied, the voltage dependence of a and 13 shifted along with EK, as is characteristic of IR channels in other cells. The steady-state voltage dependence of the gating process was well described by a Boltzmann function. The half-activation potential was on average ~7 mV negative to the observed reversal potential in all [K+]o regardless of [Mg~+]i. The activation curve was somewhat steeper when Mg-free pipette solutions were used (slope factor, 4.3 mV) than when pipettes contained 1.9 mM Mg 2+ (5.2 mV). The simplest interpretation of these data is that IR channels in bovine pulmonary artery endothelial cells have an intrinsic gating mechanism that is not due to Mg block.

“…The free ionic concentrations in relaxing and activating solutions were calculated using a computer program that solved the set of equations which described the multiple equilibria in the solutions . The values for the true affinity constants were taken from Martell and Smith (1974), except for the following (in log,u form): 10 .90 EGTA'-/Ca s+, 5.33 HEGTA'-/ Ca s+ (Kentish, 1984) ; 1 .15 HCP4-/Ca s+, 1 .30 HCP2-/Mgs+ (Smith and Alberty, 1956). The constants for equilibria involving protons were converted from concentration constants to mixed constants by the addition of 0.11 log unit (Martell and Smith, 1974) .…”

Section: Solutionsmentioning

confidence: 99%

“…The values for the true affinity constants were taken from Martell and Smith (1974), except for the following (in log,u form): 10 .90 EGTA'-/Ca s+, 5.33 HEGTA'-/ Ca s+ (Kentish, 1984) ; 1 .15 HCP4-/Ca s+, 1 .30 HCP2-/Mgs+ (Smith and Alberty, 1956). The constants for equilibria involving protons were converted from concentration constants to mixed constants by the addition of 0.11 log unit (Martell and Smith, 1974) . The calculated free concentrations of some ionic species in the solutions were: 140 mM K+ , 40 mM Na', 1 .72-2 .13 mM Mg2+, 1 nM (relaxing solution) and 2.4-190 /AM (activating solutions) Cat+, 4.2-4 .3 mM MgATP Y-, 140 mM Cl-.…”

Section: Solutionsmentioning

confidence: 99%

Some characteristics of Ca2+- regulated force production in EGTA-treated muscles from rat heart.

Kentish¹,

Jewell²

1984

McClellan and Winegrad (1980, J. Gen. Physiol., 75:283-295) have reported that in rat ventricular muscles that have reportedly been made "hyperpermeable" to small ions such as Ca2+, CaEGTA2-, and MgATP2- by a soak in EGTA, the maximum Ca2+-regulated force can be permanently increased by a short exposure to positively inotropic drugs, such as epinephrine or cAMP plus theophylline, in the presence of the detergent Triton X-100. The experiments reported here were begun as an attempt to repeat and extend this important observation. However, no evidence could be found for a potentiation of force that was not merely produced by Triton alone. In addition, the thickest muscles used (250-440 microns diameter) exhibited very low values for force per unit cross-sectional area, which suggested that either Ca2+ reached only a fraction of the myofibrils or the myofibrils were in a state of low contractility. The results of further experiments that were designed to test the permeability characteristics of these EGTA-treated muscles indicated that the movement of certain ions into these preparations was restricted, even in thin muscles (80-200 microns diameter). The rate of development of Ca2+-regulated force was slow (t1/2 approximately equal to 1-3 min), but was greatly accelerated after the muscles had been superfused with Triton X-100 (t1/2 approximately equal to 10-20 s). Removal of creatine phosphate (CP) in the presence of MgATP produced a partial rigor contracture in the EGTA-treated muscles. The results were consistent with the suggestion that the EGTA-treated muscles were permeable to some extent to Ca2+ and HCP2- ions but not to CaEGTA2- and MgATP2-. Thus, it would seem unlikely that the [Ca2+], [MgATP2-], and [Mg2+] in the immediate vicinity of the myofibrils in these preparations can be adequately controlled by the solution bathing the muscles.