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J S Fan

Publications and source records attributed to J S Fan.

At least 19 recordsLinked to original sources
PubMed2000

Kinetic effects of FPL 64176 on L-type Ca2+ channels in cardiac myocytes.

To characterize the effects of the Ca2+ channel agonist FPL 64176 on L-type Ca2+ current in isolated rat ventricular myocytes, certain of its effects were compared with those of a better known agonist, S (-) Bay K 8644. Both drugs enhance currents elicited by depolarizing pulses and enhance and slow the decay of tail currents elicited by subsequent repolarization. Both drugs shift the voltage dependence of activation and of inactivation approximately 10 mV in the negative direction, but FPL 64176 slows the rate of both activation and the decline of Ca2+ current during a depolarization, whereas Bay K 8644 accelerates the rate of current decay under the same conditions. In single channel studies in on-cell recording mode, FPL 64176 produced a great lengthening of the channel open time, produced very long openings when the channels were repolarized after a depolarizing stimulus, and had only modest effects on mean closed times and on first latency distributions. FPL 64176 and Bay K 8644 also had minimal effects on L-type channel "on" gating currents, while the "off" gating currents were slowed, particularly at positive potentials. However, the effects on gating currents were too small to account for the prolonged tails observed in FPL 64176. Once the channel is open, FPL 64176 slows transitions to closed or inactivated channel states.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed2000

Formation of a native-like beta-hairpin finger structure of a peptide from the extended PDZ domain of neuronal nitric oxide synthase in aqueous solution.

Neuronal nitric oxide synthase (nNOS) is targeted to the cell membrane via interactions of its extended PDZ domain with PDZ domains of membrane-associated proteins including PSD-95 and alpha1-syntrophin. The formation of heterodimers between the nNOS PDZ domain and the PDZ domains of nNOS-binding proteins requires a stretch of continuous amino-acid residues C-terminal to the canonical nNOS PDZ domain. In this work, we show that a 27-residue peptide comprising the C-terminal extension of the extended nNOS PDZ domain is capable of binding to PSD-95. The structure of the 27-residue peptide in aqueous solution was determined using multidimensional NMR-spectroscopic techniques. The free peptide adopts a native-like beta-hairpin finger structure in aqueous solution. The results indicate that the C-terminal extension peptide of the nNOS PDZ domain may represent a relatively independent structural unit in the mediation of the interaction between nNOS and PDZ domain-containing proteins including PSD-95 and alpha1-syntrophin.

Amino Acid Sequence↗
PubMed1999

One calcium ion may suffice to open the tetrameric cardiac ryanodine receptor in rat ventricular myocytes.

1. The release of Ca2+ from sarcoplasmic reticulum in response to Ca2+ entering through L-type Ca2+ channels was studied in isolated voltage clamped rat ventricular myocytes at room temperature using the fluorescent Ca2+ indicators fluo-3 and Oregon Green 488 Bapta 5N. 2. Depolarizations to positive potentials elicited fluo-3 Ca2+ transients with rates of rise that were linearly related to the magnitude of the peak measured Ca2+ current in the presence of Cs+-containing pipette solutions. 3. Further experiments utilizing prepulses to preactivate a constant number of channels also revealed a linear relationship between the Ca2+ transient rate of rise and the magnitude of entering Ca2+ current at positive potentials. Under these conditions as well, the maximal rates of rise of global myoplasmic Ca2+ transients were due primarily to Ca2+ release from the sarcoplasmic reticulum as revealed by effects of ryanodine and caffeine on the Ca2+ transients. Using such prepulses, linearity between the Ca2+ transient rate of rise and the magnitude of the peak Ca2+ current was found under a variety of pulse protocols. 4. Using one such pulse protocol, linearity between the Ca2+ transient rate of rise and the magnitude of the peak Ca2+ current was also found when Ca2+ currents assessed at one potential were reduced in magnitude during the onset of block by application of Co2+. Using the same pulse protocol, linearity between the Ca2+ transient rate of rise and the magnitude of the peak Ca2+ current was also found when use of Cs+ was avoided by blocking K+ currents with extracellular TEA and 4-aminopyridine. Linearity in the relationship between the Ca2+ transient rate of rise and the magnitude of the peak Ca2+ current was also found when Ca2+ transients were measured using the low affinity Ca2+ indicator Oregon Green 488 Bapta 5N in place of fluo-3. 5. These results appear to indicate that the cardiac ryanodine receptor is capable of being activated by only one calcium ion. Alternative interpretations of the data are discussed.

Animals↗
PubMed1999

Effects of outer mouth mutations on hERG channel function: a comparison with similar mutations in the Shaker channel.

The fast-inactivation process in the hERG channel can be affected by mutations in the pore or S6 domain, similar to the C-type inactivation in the Shaker channel. However, differences in the kinetics and voltage dependence of inactivation between these two channels suggest that different structural determinants may be involved. To explore this possibility, we mutated a serine in the outer mouth region of hERG (S631) to residues of different physicochemical properties and compared the resulting changes in the channel's inactivation process with those resulting from mutations of an equivalent position in the Shaker channel (T449). The most dramatic differences are seen when this position is occupied by a charged residue: S631K and S631E disrupted C-type inactivation in hERG, whereas T449K and T449E facilitate C-type inactivation in Shaker. S631K and S631E also disrupted the K selectivity of hERG pore, a change not seen in T449K or T449E of Shaker. To further study why there are such differences, we replaced S631 with cysteine. This allowed us to manipulate the properties of thiol groups at position 631 and correlate side-chain properties here with changes in channel function. S631C behaved like the wild-type channel when the thiol groups were in the reduced state. Oxidizing thiol groups with H2O2 or modifying them with MTSET or MTSES disrupted C-type inactivation and K selectivity, similar to the phenotype of S631K and S631E. The same thiol-modifying maneuvers did not affect the wild-type channel function. Our results suggest differences in the outer mouth structure between hERG and Shaker, and we propose a "molecular spring" hypothesis to explain these differences.

Amino Acid Sequence↗
PubMed1999

Heterogeneous changes in K currents in rat ventricles three days after myocardial infarction.

OBJECTIVE: After coronary artery occlusion, surviving myocardium in and around the infarct zone plays an important role in arrhythmogenesis. Understanding the mechanisms for derangements in cardiac electrical activity at the cellular and molecular levels is important for the design of effective therapeutic strategies. METHODS: To provide part of that understanding, we studied changes in K channel function and expression in rat ventricular myocardium three days after occluding the left major coronary artery. The epicardium and endocardium of infarcted region in the left ventricle and the free wall of right ventricle were separated for myocyte isolation, followed by whole-cell voltage clamp studies. Myocytes were also isolated from corresponding regions of control and sham-operated hearts and studied under the same conditions. RESULTS: We found that the transient outward (Ito), delayed rectifier (IK) and inward rectifier (IKI) currents have different distribution patterns in normal rat ventricular myocardium. Sham-operation did not affect any of these K currents in left ventricular myocytes, but coronary artery occlusion caused a reduction of all three. For Ito and IKI the reduction was greater in epicardial than in endocardial myocytes, but IK was reduced equally in these two cell groups. Unexpectedly, Ito and IK as well as cell capacitance were increased in right ventricular myocytes from infarcted as well as sham-operated hearts. Western blot analysis indicated that the level of Kv4 channel proteins (Kv4.2 + Kv4.3) was reduced in infarcted left ventricular myocardium, consistent with the reduction in Ito. CONCLUSION: Our data suggest that the distribution of K channels and changes in them induced by coronary artery occlusion are heterogeneous in ventricular myocardium. Understanding the molecular mechanisms for this heterogeneity and its implications in arrhythmogenesis poses a challenge in designing effective antiarrhythmic therapy for myocardial infarction patients.

Animals↗
PubMed1999

Use-dependent 'agonist' effect of azimilide on the HERG channel.

Azimilide (AZ) is a class III antiarrhythmic drug that has voltage-dependent dual effects on the HERG channel: 1) increasing current amplitude at low-voltage depolarization (agonist effect), and 2) suppressing current at more depolarized voltages (antagonist effect). We examined the mechanism for the agonist effect of AZ on HERG expressed in Xenopus oocytes. The agonist effect resulted from an AZ-induced 'prepulse potentiation: a strong depolarization prepulse increased the rate and degree of channel activation induced by subsequent depolarization to -50 or -40 mV. The potentiated state decayed slowly in an exponential fashion (time constant, 60-80 s). Degrees of potentiation were proportional to degrees of channel activation during prepulses; hence, the agonist effect of AZ was use dependent. AZ exerted its agonist effect from outside the cell membrane, and the effect did not depend on intracellular G-protein or protein kinase activity. Mutations made in the outer mouth or an extracellular loop connecting the S5 and P regions of HERG, which could hinder or modify conformational changes in the pore region during membrane depolarization, reduced or abolished AZ-induced prepulse potentiation. Importantly, these same mutations also increased the rate and degree of channel activation in the negative voltage range, and the degree of change in the activation properties was inversely correlated with the degree of AZ-induced prepulse potentiation. We propose that conformational changes in the outer mouth and neighboring extracellular domain of HERG during membrane depolarization can affect the process of channel activation. In the presence of AZ, channel activation allowed drug modification of these conformational changes, which subsequently facilitated HERG activation by low-voltage depolarization.

Animals↗
PubMed1998

A peptide corresponding to residues Asp177 to Asn208 of human cyclin A forms an alpha-helix.

Cyclins are essential activators of eukaryotic cell cycle-regulating enzymes called cyclin-dependent kinases (CDKs). The binding of cyclins to CDKs is mediated by a structural motif comprising a five-helix bundle called the cyclin fold and an additional helix (the N-terminal alpha-helix) located N-terminal to the cyclin fold. In this work, we examine, using CD and NMR spectroscopy, the structure of a 32-residue synthetic peptide derived from the segment (Asp177 to Asn208) corresponding to the N-terminal alpha-helix of human cyclin A. CD spectroscopic analysis of the peptide revealed that trifluoroethanol (TFE) can induce the peptide to assume a stable alpha-helix conformation. Two-dimensional 1H NMR spectroscopy showed that the alpha-helix is formed by the Asp181 to Cys193 segment of the peptide. The alpha-helical structure of the peptide in the TFE/H2O cosolvent was found to be identical to that in the crystal structure of intact cyclin A. Taken together, these results suggest that the N-terminal alpha-helix of cyclins may exist as an independent structural unit that plays essential functional roles in activating CDKs.

Amino Acid Sequence↗
PubMed1998

Protein inhibitor of neuronal nitric-oxide synthase, PIN, binds to a 17-amino acid residue fragment of the enzyme.

Neuronal nitric-oxide synthase (nNOS) is the primary nitric oxide (NO) regulator in neurons. The activity of the enzyme is inhibited by a protein inhibitor called PIN. We were able to purify large quantities of PIN overexpressed in bacterial cells. Analytical ultracentrifugation and chemical cross-linking studies showed that PIN exists as a monomer at low concentrations. The protein forms a high order aggregate at elevated concentrations. We have shown, using NMR spectroscopy, that the previously identified PIN-binding domain (PINB) of nNOS (residues 161-245) adopts a random coil structure in solution. By titrating 15N-labeled PINB with unlabeled PIN, the PIN-binding region of nNOS was precisely mapped to a 17-residue peptide fragment from Met-228 to His-244 of nNOS. NMR titration experiments also showed that PIN binds to nNOS with a 1:2 stoichiometry. A synthetic peptide corresponding to the identified PIN-binding region of nNOS was used to study the interaction between PIN and nNOS in detail. The functional implications of the results obtained from this study are discussed.

Amino Acid Sequence↗
PubMed1998

Perforated patch recording with beta-escin.

Perforated patch recording with nystatin, amphotericin B and gramicidin can be more difficult in the hands of some investigators than others. In addition, it is difficult to introduce low molecular weight substances such as dyes into the cytoplasm in such experiments. We have determined that beta-escin represents a convenient, easy-to-use alternative to less water-soluble ionophores.

Amphotericin B↗
PubMed1998

The relationship between strength and function in females with juvenile rheumatoid arthritis.

OBJECTIVE: To describe the relationship between isokinetic concentric and eccentric quadriceps strength and function in females with juvenile rheumatoid arthritis (JRA). METHODS: Twenty girls with JRA aged 6 to 16 years participated in the study. Function and strength were measured on one occasion. Isokinetic concentric and eccentric quadriceps torque was measured on the Kin-Com dynamometer. Function was measured with the Childhood Health Assessment Questionnaire (CHAQ) and the Canada Fitness Award 50 m run. Pain and joint count were taken as measures of disease activity. Pain was assessed on a visual analog scale through parent- and self-report. Correlations between function, standardized torque values, pain, and joint count were determined. Stepwise multiple regression analyses with function as the predicted variable were performed. RESULTS: Correlations between standardized torque and CHAQ scores were moderate (r = -0.48, p = 0.03 for concentric torque and r = -0.43, p = 0.06 for eccentric). Correlations between torque and run scores were also moderate (r = -0.48, p = 0.03 for concentric, r = -0.31, p = 0.19 for eccentric). Pain and total joint count were the best predictors of CHAQ score (r2 = 0.80), while concentric torque was the best predictor of running ability (r2 = 0.23). CONCLUSION: Quadriceps torque was moderately correlated to function in girls with JRA. Function as determined on the CHAQ was best predicted from measures of disease activity, while function as determined on the 50 m run was best predicted from concentric quadriceps torque. Further research is required to determine a cause-effect relationship between strength and function in girls with JRA.

Adolescent↗
PubMed1997

Effects of elevating [Na]i on membrane currents of canine ventricular myocytes: role of intracellular Ca ions.

OBJECTIVES: Our first objective was to study how elevating [Na]i can modify the background membrane conductance in canine ventricular myocytes (CVM). In particular, we wanted to find evidence for a Nai-activated K current (IK,Na) in these cells. The second objective was to compare the effects of elevating [Na]i on membrane currents without and with intracellular Ca buffering. METHODS: Whole-cell currents were recorded and [Na]i was elevated either by using a pipette perfusion device that allowed [Na] in the pipette solution to be varied (from 0 to 50 mM), or by 50 microM ouabain. RESULTS: Although an outward current attributable to IK,Na was confirmed in guinea-pig ventricular myocytes (GPVM) under our recording conditions, no such current was seen in 29 CVM examined. With Cai buffering, the main effect of elevating [Na]i on CVM was an increase in inward current around and negative to the resting membrane potential. Based on the dependence of this Nai-induced inward current on K ions and its pharmacological properties, especially the effects of low concentrations of external Ba ions (< or = 5 microM) at strongly hyperpolarized voltages, we hypothesize that this current was carried by extracellular K ions through the inward rectifier (IK1) channels that had been modified by the high level of [Na]i. With Cai buffering, elevating [Na]i by ouabain had few or no effects on the L-type Ca channel current (ICa) or the slow delayed rectifier current (IKs). Without Cai buffering, ouabain induced a rapid reduction of both currents along with an increase in a time-independent outward current at voltages positive to -60 mV. CONCLUSION: Our data suggest that there are species variations in K channel expression and/or K channel modulation by intracellular Na ions. Furthermore, intracellular Ca ions play a crucial role in mediating the effects of Nai loading on membrane currents in canine ventricular myocytes.

Animals↗
PubMed1993

[Effects of panaxatriol saponins (PTS) isolated from panax notoginseng on the action potential and delayed rectifier current (Ix) in sheep cardiac Purkinje fibers].

The electrophysiological effects of PTS in sheep cardiac Purkinje fibers were studied. PTS was shown to increase the duration of action potential (APD30, APD50 and APD90) at the concentrations of 2.5 micrograms/ml and 5.0 micrograms/ml. However, the amplitude of action potential (APA) remained unchanged. The result of using double microelectrode voltage clamp method showed that PTS (1.25-10.0 micrograms/ml) depressed the delayed (outward) rectifier current (Ix) in time- and dose-dependent manners, when the holding potential was held at +20 mV, the command potential was held at +10 mV, 0.2 Hz and the clamping time at 1-1.5 s. It may be concluded that the effect of PTS on APD is mainly related to blocking the delayed rectifier potassium channel.

Action Potentials↗
PubMed1983

Inter-relationships between sodium and potassium intake and the blood pressure effects of ACTH in sheep.

These studies examine the effect of sodium (Na) and potassium (K) intake on the pressor and metabolic actions of ACTH (20 micrograms/kg/day) in sheep. After 21 days on each of five regimens in which Na and K intake varied from 0 to 100 mmol/day, no simple relationship between Na and K intake and blood pressure was found. After five days of ACTH treatment, mean arterial pressure (MAP) rose + 5 mmHg in sheep on 0 mmol Na, 0 mmol K (expressed as 0 Na 0 K); + 13 mmHg on 10 Na 100 K; + 5 mmHg on 0 Na 100 K; + 20 mmHg on 100 Na 0 K and + 24 mmHg on 100 Na 100 K. Plasma [K] was unchanged by ACTH on 0 Na 0 K but fell in sheep on the other electrolyte regimens. Water intake increased with ACTH on all regimens except 100 Na 0 K. Blood aldosterone concentration was high in sheep maintained on 0 Na regimens but lower after five days of ACTH treatment in all groups. Blood cortisol and corticosterone concentrations increased with ACTH on all regimens studied.

Adrenocorticotropic Hormone↗
PubMed1981

Effect of adrenocortical steroids and their structural analogues on blood pressure in sheep.

1. A model of adrenocortical steroid-induced hypertension based on the effects of ACTH administration has been developed in sheep. The present studies examine the effects of a number of different steroid hormones on blood pressure to investigate their structure-activity relationships. 2. Infusion of the major ovine adrenal steroid hormones (combined steroid infusion of cortisol, corticosterone, 11-deoxycortisol, aldosterone, deoxycorticosterone) reproduced the metabolic but not the blood pressure effects of ACTH. 3. Addition of 17 alpha, 20 alpha-dihydroxyprogesterone and 17 alpha-hydroxyprogesterone, at rates appropriate for conditions of ACTH stimulation, to the combined steroid infusion reproduced both the blood pressure and metabolic effects of ACTH. 4. 17 alpha-20 beta-Dihydroxyprogesterone, 20 beta-dihydroxy-11-deoxycortisol and 20 beta-hydroxycortisol all had additional hypertensive activity when given with combined steroid infusion, but 16 alpha-hydroxyprogesterone, 11 beta, 17 alpha-dihydroxyprogesterone, 20 alpha-hydroxyprogesterone and 20 beta-hydroxyprogesterone did not. 5. These studies support the concept of a new class of steroid hormone action in blood pressure regulation.

Adrenal Cortex Hormones↗
PubMed1980

ACTH-elicited sodium appetite in sheep.

Intramuscular injections of long-acting synthetic ACTH (45 U twice daily for 5 days) caused a large increase in the intake of 0.5 M NaCl in sheep. Mean Na intake of the sheep on the last 3 days of treatment approximated 50% of their total extracellular fluid Na. The mineral appetite was specific for NaCl. Intakes of 0.5 M KCl or 0.25 M CaCl2 were not significantly altered. The enhanced appetite for Na induced by ACTH appeared to precede any increase in urinary Na excretion. ACTH treatment was ineffective in adrenalectomized sheep. However, an infusion into adrenalectomized sheep of a combination of adrenal steroid hormones (including aldosterone, deoxycorticosterone, 11-deoxycortisol, cortisol, and corticosterone) that contrived blood levels similar to those, obtained with ACTH treatment in normal sheep did induce Na appetite. Thus, ACTH induces a specific, adrenal-steroid hormone-dependent Na appetite in sheep.

Adrenocorticotropic Hormone↗
PubMed1978

Exaggerated natriuresis in ACTH hypertension in sheep.

Adrenocorticotrophic hormone (ACTH) administration to sheep (100 IU/day) produces a sustained increase in arterial pressure within 24 h. The effect of ACTH on excretion of an intravenous saline load was tested in 8 adult cross-bred Merino ewes. A significant diuretic response to saline loading was found after 24 h but no increased natriuresis. On days 3 and 6, diuretic and natriuretic responses were both significantly increased. Steroid-induced sodium retention may have abolished the natriuretic effect on day 1.

Adrenocorticotropic Hormone↗