Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Potassium currents in ventricular myocytes from genetically diabetic rats KATSUHARU TSUCHIDA AND HIROSHI WATAJIMA Research Center, Taisho Pharmaceutical Company, Ohmiya, Saitama 330, Japan cardiomyocytes; transient outward current that patients with diabetes mellitus have increased risk of mortality from cardiac failure that cannot simply be explained in terms of atherosclerosis, hyperlipidemia, or hypertension. Many studies have been performed to investigate the cardiac function of diabetic animals, and a variety of mechanical, electrophysiological, and biochemical abnormalities have been identified in the myocardium of diabetic animal models (7). Chemical-induced diabetic models, especially the streptozotocin (STZ)-induced diabetic model, are frequently used as a model for insulindependent diabetes mellitus (IDDM) to investigate these abnormalities. With use of this animal model, contractile abnormalities have been reported (7), and this dysfunction has been related to biochemical changes such as the altered distribution of myosin isozymes (4); a decrease in uptake of Ca21 into sarcoplasmic reticulum (6) and sarcolemma (9); decreased activity of Na1-K1-adenosinetriphosphatase (14), Na1-H1 exchange (15), and Na1-Ca21 exchange (16) in the sarcolemmal membrane; and Ca21 binding capacity (20). Abnormalities of electrophysiological phenomena have also been demonstrated in STZ-induced diabetic rats. Ventricular and atrial action potentials were prolonged IT HAS LONG BEEN KNOWN when the measurement was done for a relatively longer term after STZ injection (1, 15). The L-type calcium current was not altered from 6 days to 2 mo (12, 17) but was decreased from 5 to 7 mo (28), whereas the transient outward potassium current was decreased significantly from 6 days to 7 mo after STZ injection (12, 17, 24, 28). Our previous study using genetically diabetic rats (WBN/Kob) demonstrated that the ventricular action potential duration is prolonged (25). STZ-treated rats closely resemble the IDDM model, except in a special case (23), whereas WBN/Kob rats show some similarities with the non-insulin-dependent diabetes mellitus (NIDDM) model. That is, male WBN/ Kob rats have been reported to develop hyperglycemia, glycosuria, polyuria, and glucose tolerance from ,9 mo of age, with a gradual and moderate decrease in serum insulin levels (19). The rats survive without the administration of insulin, in contrast to Bio-Breeding rats, a genetic model of IDDM. The pathological process progresses slowly in WBN/Kob rats. Much clinical attention has been paid to NIDDM-induced cardiomyopathy (7, 23). We have previously reported that the L-type Ca21 channel did not alter significantly, although the response of the L-type Ca21 channel to b-adrenergic stimulation decreased in WBN/Kob rats (25). In this study, we first examined changes in the potassium currents, especially the transient outward potassium current, in this strain of genetically diabetic rats. MATERIALS AND METHODS Animal model of diabetes. A particular strain of Wistarderived WBN rats has been maintained at the Institute of Pathology, Bonn University (Bonn, Germany). Several of these animals were brought to Japan by Dr. O. Kobori in 1976 and became known as WBN/Kob rats. A colony of these rats is currently being maintained at the Shizuoka Laboratory Animal Center (Shizuoka, Japan). Male WBN/Kob rats of 17 to 19 mo of age and age-matched Wistar rats (purchased from the Shizuoka Laboratory Animal Center) were used. The main clinical sign of diabetes, glucosuria, was evidently detected at ,14 mo of age, after a marked glucose intolerance at 12 mo. Thereafter, some animals developed hyperlipidemia and gradual emaciation (19, 26). After purchase at 6 mo and until they were used in the experiment, all rats were maintained at the Taisho Pharmaceutical Animal Laboratory and were fed standard rat chow. Cell preparation. Single ventricular cells were isolated according to the methods previously reported (25). Briefly, the rat was anesthetized with pentobarbital sodium (50 mg/kg ip), and the heart was rapidly excised and attached to a Langendorff perfusion apparatus. The heart was then retrogradely perfused for 2–3 min with nominally calcium-free Krebs-Henseleit solution equilibrated with 95% O2-5% CO2 at 36°C. The Krebs-Henseleit solution contained (in mM) 130 NaCl, 4.8 KCl, 1.2 MgSO4, 1.1 NaH2PO4, 25 NaHCO3, and 0193-1849/97 $5.00 Copyright r 1997 the American Physiological Society E695 Downloaded from http://ajpendo.physiology.org/ by 10.220.33.1 on May 2, 2017 Tsuchida, Katsuharu, and Hiroshi Watajima. Potassium currents in ventricular myocytes from genetically diabetic rats. Am. J. Physiol. 273 (Endocrinol. Metab. 36): E695–E700, 1997.—Our previous study demonstrated the longer duration of action potential in ventricular myocytes from genetically diabetic WBN/Kob rats without change in calcium channel density compared with age-matched controls [Tsuchida, K., H. Watajima, and S. Otamo. Am. J. Physiol. 267 (Heart Circ. Physiol. 36): H2280–H2289, 1994]. In the present study we examined the alteration of potassium currents, especially transient outward current, in ventricular myocytes of genetically diabetic WBN/Kob rats. WBN/Kob rats gradually develop hyperglycemia with aging and show some similarity to non-insulin-dependent diabetes mellitus models, which differ from the insulin-dependent streptozotocin-treated rat model. The density of the intracellular calcium ion-independent transient outward current (Ito ) from 17- to 19-mo diabetic rat myocytes was significantly smaller than that from age-matched control rat myocytes. In addition, the density of Ito from 17- to 19-mo rat myocytes was significantly less than that from 2-mo rat myocytes, suggesting that aging-induced alteration of Ito was accelerated by the diabetic state. The steady-state inactivation curves of Ito, the recovery from Ito inactivation, and the other outward currents were not significantly altered between diabetic myocytes and agematched control myocytes. In conclusion, the prolonged duration of action potential from genetically diabetic rat myocytes is mainly due to the depressed Ito. E696 POTASSIUM CURRENTS IN GENETICALLY DIABETIC HEARTS Fig. 1. Representative outward currents recorded in single ventricular myocytes isolated from diabetic (17– 19 mo) and age-matched control rat heart and in young adult (2 mo) normal rat ventricular myocytes. Currents were elicited by applying a 300-ms depolarizing step in 10-mV increments to 170 mV from a holding potential of 270 mV every 5 s. Effects of 5 mM 4-aminopyridine (4-AP) on the current elicited by a test pulse to 170 mV from 270 mV are also represented. Six families of current tracings are from different myocytes. Total cell capacitances (pF) are as follows: right (R) 170, left (L) 132 in 17- to 19-mo control rats; R 153, L 186 in 17- to 19-mo diabetic rats; R 117, L 144 in 2-mo rats. tetrodotoxin (TTX)-sensitive Na1 current, the Na1-activated K1 current, and ICa. The composition of Na1-free solution was (in mM) 135 choline chloride, 5.4 KCl, 1 MgCl2, 1.8 CaCl2, 10 HEPES, 10 glucose, and 3 CoCl2 (pH 7.4 with KOH). In some experiments, 4-aminopyridine (4-AP; Sigma Chemical, St. Louis, MO) was used to block Ito, and TTX (Sankyo, Tokyo, Japan) was used to block sodium ion current (INa). Concerning the determination of cell membrane capacitance, the procedure described below was used. Just after the patch was broken, pulses with a duration of 50 ms, from 260 to 262 mV, were applied to the cell. The exponential components of the decaying current were determined. Two time constants were obtained corresponding to the electrode and membrane capacitance. The time constant of the electrode was ,0.02 ms, and the time constant of the membrane capacitance was just under 2 ms. After the electrode capacitance was compensated for, the capacitance of the membrane was calculated according to the equation Cm 5 Tc · Io /Em [1 2 (I/Io)] where Cm is the membrane capacitance, Tc is the time constant of the membrane capacitance, Io is the maximum capacitance current value, Em is the amplitude of the voltage step, and I is the steady-state current. The series resistance (Rs ) was calulated as Rs 5 Em /Io which ranged from ,4 to 10 MV. In some cases, membrane capacitance and series resistance were determined electronically. Then membrane capacitance was compensated for, and series resistance was reduced maximally for the voltageclamp experiments to examine the membrane currents. Downloaded from http://ajpendo.physiology.org/ by 10.220.33.1 on May 2, 2017 12.5 glucose. Enzymatic digestion was achieved by recirculating the perfusion apparatus with the calcium-free KrebsHenseleit solution containing 40–170 U/ml collagenase (Yakult, Tokyo, Japan). The perfusion pressure was maintained at ,80 mmHg. Enzymatic perfusion was stopped after 15–30 min. The heart was then washed with Kraftbrühe (KB) solution containing (in mM) 70 L-glutamic acid, 5 KCl, 20 taurine, 5 KH2PO4, 11 glucose, 5 N-2-hydroxyethylpiperazineN8-2-ethanesulfonic acid (HEPES), and 0.5 ethylene glycolbis (b-aminoethyl ether)-N,N,N8,N8-tetraacetic acid (EGTA) free acid. The ventricle was separated from the whole heart and minced into small pieces. Cells were filtered and stored in KB solution at 4°C before being used in the electrophysiological experiment. After storage for 3–6 h in KB solution, the cells were taken into the experimental chamber, which was perfused with Ca21-containing Tyrode solution when the electrophysiological experiment was performed. Electrophysiological recording. Some cells were transferred to a recording chamber (0.3 ml vol) placed on the stage of an inverted microscope (Diaphot TMD, Nikon, Tokyo, Japan), and the chamber was perfused at a constant rate of 1–2 ml/min. Membrane currents were recorded using the whole cell patch-clamp method described by Hamill et al. (8) by use of a patch-clamp amplifier (CEZ-2300, Nihon Kohden) connected to pClamp software program (Axon Instruments, Burlingame, CA) or an Atari Mega ST4-operated EPC-9 patch-clamp system (Heka, Lambrecht, Germany). Patch pipettes (2–4 MV) were fabricated using a puller (PP-83, Narishige, Tokyo) and were heat polished (MF-83, Narishige). To record transient outward current (Ito ), we filled the pipettes with solution containing (in mM) 140 KCl, 2 MgCl2, 1 CaCl2, 10 HEPES, 11 EGTA, and 5 Na2ATP (pH ajusted to 7.2 with KOH). The extracellular mediums were standard Tyrode solution containing Co21, to eliminate a calcium ion current (ICa ), or a Na1-free Co21-containing solution to eliminate the POTASSIUM CURRENTS IN GENETICALLY DIABETIC HEARTS E697 Data analysis. Statistical significance was determined with the Student’s t-test for unpaired data or with the repeated-measures analysis of variance test for multiple comparison. A value of P , 0.05 was considered significant. RESULTS AND DISCUSSION Fig. 2. Current-voltage relationships of transient outward current (Ito ) and terminal components of the outward current in ventricular myocytes from diabetic (17–19 mo) and age-matched control rats, and from young adult (2 mo) rats. Amplitude of Ito was measured as the difference in current amplitude between the peak current amplitude and the terminal current amplitude at the end of 300-ms test pulses. A: Ito; B: terminal current. r, Diabetic myocytes (n 5 23); s, age-matched control myocytes (n 5 22); k, young adult myocytes (n 5 14). Inset a: voltage-clamp protocol; inset b: 4-AP-sensitive current induced by a 300-ms test pulse to 170 mV from 270 mV. Control, n 5 16; WBN/Kob, n 5 16. Values are means 6 SE. * P , 0.05 vs. age-matched control (17–19 mo) for diabetic rat myocytes, and 1 P , 0.05 vs. young adult (2 mo) rat for aged (17 – 19 mo) rat myocytes at each membrane potential (Student’s t-test). A supplemental explanation: repeated-measures analysis of variance test showed that current (Ito )-voltage relationships were significantly (P , 0.05) different between diabetic rats and age-matched control rats. Current (Ito )voltage relationships were also significantly (P , 0.05) different between 2-mo rats and 17- to 19-mo rats. consistent with findings reported in STZ-treated diabetic rats (12, 17, 28). We used aged rats of 17–19 mo as controls, because WBN/Kob rats gradually develop hyperglycemia with aging. We examined the influence of aging on Ito as well. Wei et al. (29) demonstrated that Downloaded from http://ajpendo.physiology.org/ by 10.220.33.1 on May 2, 2017 Plasma glucose level and clinical state. Plasma glucose levels were 558 6 16 (SE) mg/dl with 17- to 19-mo WBN/Kob rats (n 5 34), whereas plasma glucose levels of age-matched control Wistar rats were within the range of ,110–140 mg/dl. Almost all WBN/Kob rats showed glucosuria and ruffled hair including transient alopecia and seemed sluggish at 17–19 mo. Effects of genetic diabetes on Ito. Outward currents were recorded after blocking ICa with 3 mM Co21 or INa with 30–50 mM TTX, or changing extracellular NaCl with choline chloride and the intracellular Ca21activated current with 11 mM EGTA added in the pipettes. From a holding potential of 270 mV, depolarizing pulses in 10-mV increments were applied from 260 mV up to 170 mV. Ito was elicited by depolarizing pulses more positive than 220 mV in both normal and diabetic ventricular myocytes. Application of 5 mM 4-AP suppressed Ito, and the 4-AP-insensitive component of the outward current remained. Figure 1 shows typical currents obtained from diabetic (17–19 mo), age-matched control (17–19 mo), and young (2-mo) rat ventricular myocytes. Ito consists of 4-AP-sensitive and -insensitive components. The 4-AP-insensitive Ito is reported to be activated by increased intracellular calcium concentration ([Ca21]i ). Because we used a pipette solution containing a relatively high concentration of EGTA (11 mM), the [Ca21]i-dependent activation of Ito could be ruled out, thereby leaving only one component of Ito, namely, the 4-AP-sensitive component shown in Fig. 1. Hereafter, we will refer only to this component of Ito. The amplitude of Ito was expressed as the difference in current between the peak current amplitude and the terminal component of the current at the end of depolarizing test pulses of 300-ms duration. The amplitude of Ito is shown in Fig. 2A. The current density of the difference in current between peak and 300-ms terminal currents was significantly less in diabetic ventricular myocytes than in the age-matched control myocytes. Apkon and Nerbonne (2) described a slowly activating and inactivating K1 current (IK in their terminology) in rat ventricular myocytes, which was sensitive to external tetraethylammonium but not to 4-AP. Thus the terminal component at the end of 300-ms pulses is considered to consist of two different currents. One is the residual component of the slowly inactivating Ito at the end of 300-ms pulses, and the other is the 4-APinsensitive outward current. To determine the total Ito as the 4-AP sensitive current, the difference in current between the absence and the presence of 5 mM 4-AP was obtained and shown in the inset of Fig. 2A. The density of the 4-AP-sensitive current was also significantly less in the diabetic ventricular myocytes than in the age-matched control myocytes. The decrease of Ito density in the genetically diabetic WBN/Kob rats is E698 POTASSIUM CURRENTS IN GENETICALLY DIABETIC HEARTS the action potential duration in the senescent (24-mo) rat ventricular myocytes was longer than that in the young adult (7-mo) rat myocytes. Other authors have reported similar changes of action potential duration (3). Walker et al. (27) demonstrated that Ito was depressed more markedly in the cardiac cells of the aged (24- to 25-mo) rat than in those of young adult (2- to 3-mo) rats. The current density of Ito from 17- to 19-mo rat myocytes was significantly less than that from 2-mo rat myocytes (Fig. 2A). The decay process of Ito seemed to be fitted with biexponential curves with fast and slow time constants (tf and ts, respectively). Because the pulse duration was too short to determine ts correctly, we have presented only tf. At 170 mV, tf was 32.8 6 2.9 ms (n 5 21) in 17- to 19-mo control, 32.3 6 1.2 ms (n 5 19) in 17- to 19-mo diabetic, and 33.4 6 1.5 ms (n 5 14) in 2-mo normal rats. All values were not significantly different. Possible values of ts seemed roughly ten times longer than those of tf. The terminal current amplitude at the end of 300-ms pulses is shown in Fig. 2B. The terminal component was not altered in WBN/Kob rat myocytes compared with that in the age-matched control rat myocytes. Jourdon and Feuvray (12) and Wang et al. (28) described the significant decrease in this time-independent outward current in the STZ-treated rat myocytes. The present result is different from their results. In addition, the current amplitude of the terminal component was not altered between examinations of 17- to 19-mo rats and 2-mo rat myocytes (Fig. 2B). Modification of inactivation kinetics of Ito. In this series of experiments, Ito was also expressed as the change in current between the peak amplitude and the terminal component of the current. The steady-state inactivation kinetics of Ito were compared between the diabetic myocytes and the age-matched control myocytes. A test pulse to 170 mV (duration 300 ms) was preceded by 1-s conditioning prepulses to various potentials (from 280 mV to 0 mV). The relative amplitude of Ito (normalized by taking the value at 280 mV as unity) was plotted against conditioning potentials, and the data were fitted by the following Boltzmann distribution function with a least squares method I/Imax 5 1 / [1 1 e(Vm2V0.5)/k] where I/Imax is the relative amplitude of Ito, Vm is the conditioning voltage, V0.5 is the voltage of half-inactivation, and k is the slope factor. The values of V0.5 and k were 243.9 6 2.1 and 7.7 6 0.54 mV for age-matched control myocytes, and 247.3 6 3.1 and 6.9 6 0.38 mV for diabetic myocytes, respectively (Fig. 3). The value of V0.5 was not altered significantly between the two groups. The value of k was not significantly different between the two groups, either. Furthermore, the values of V0.5 and k were 251.6 6 2.9 and 6.6 6 0.65 mV for 2-mo rat myocytes. The V0.5 value in 2-mo rat myocytes Fig. 5. Inward rectifying potassium current density in ventricular myocytes from diabetic and age-matched (17–19 mo) control and young adult (2 mo) rats. Steady-state inward current measured at end of 1-s duration of hyperpolarizing test pulses at 10 mV increased voltage steps from a holding potential of 270 mV to 210 up to 2110 mV. r, Diabetic (n 5 15); s, age-matched (17–19 mo) control (n 5 13); k, young adult (2 mo) (n 5 10) myocytes. Values are means 6 SE. Inset: voltage-clamp protocol. Downloaded from http://ajpendo.physiology.org/ by 10.220.33.1 on May 2, 2017 Fig. 3. Steady-state inactivation curves of Ito in diabetic (17–19 mo) and age-matched controls and young adult (2 mo) rat myocytes. I/Imax, relative amplitude of Ito. r, Diabetic (n 5 18) myocytes; s, controls (n 5 16); k, young adult myocytes (n 5 10). Values are means 6 SE. Inset: voltage-clamp protocol. Fig. 4. Time course of recovery from inactivation of Ito in ventricular myocytes from diabetic (n 5 18), age-matched (17–19 mo) controls (n 5 17), and young adult (2 mo) myocytes (n 5 12). Double pulses (each duration 300 ms) from 270 mV to 170 mV were applied at varying interpulse intervals (from 10 ms to 1 s) every 10 s. r, Diabetic myocytes; s, age-matched control myocytes; k, young adult myocytes. Values are means 6 SE. Inset: voltage-clamp protocol. POTASSIUM CURRENTS IN GENETICALLY DIABETIC HEARTS understanding the pathophysiology of the diabetic heart more thoroughly. We previously demonstrated that the L-type Ca21 current was not altered in 19-mo WBN/Kob rat ventricular myocytes in vitro under the condition of the inhibition of the potassium current by use of the whole-cell patch-clamp technique (25). However, the in situ ventricular myocytes exist in circumstances without any artificial restriction of the potassium current. The action potential duration of rat ventricular cells is short enough to allow maximum Ca21 influx from extracellular medium, so that the lengthening of the action potential duration due to the inhibition of Ito may lead to increased Ca21 influx and subsequent enhancement of Ca21 release from intracellular stores, resulting in some compensatory effects on the decreased contractile force of diabetic myocardium (10, 21). Our previous study (25) indicated that WBN/Kob rats demonstrated decreased contractile force in situ. Thus the compensation resulting from lengthening the action potential duration was not considered to be sufficient for many reasons (see the introductory section of this paper) other than the decreased ICa being responsible for the decreased contractile force in WBN/Kob rats. Recently, Xu et al. (30) demonstrated that the decreased Ito density may be caused by the decrease in cellular glucose metabolism in ventricular myocytes of short-term (14 days to 1 mo) STZ-treated rats (30). Such biochemical changes may be involved in downregulating Ito in genetic NIDDM myocytes. The lower density of the Ito observed in the diabetic ventricular myocytes may result from a decrease in the number of channels. Concerning a molecular basis of the decreased Ito in diabetic myocytes, because Dixon et al. (5) demonstrated that both the ventricular potassium (Kv) 4.2 and Kv 4.3 channels were likely to contribute to the Ito in rat heart, the alteration of the expression of these genes may account for the altered Ito density in WBN/ Kob rats. The molecular alterations in the channel proteins remain to be elucidated as an interesting problem. Address for reprint requests: K. Tsuchida, Research Center, Taisho Pharmaceutical Co., Ltd., 1–403 Yoshino-cho, Ohmiya, Saitama 330, Japan. Received 25 February 1997; accepted in final form 4 June 1997. REFERENCES 1. Aomine, M., S. Nobe, and M. Arita. Increased susceptibility to hypoxia of prolonged action potential duration in ventricular papillary muscles from diabetic rats. Diabetes 39: 1485–1489, 1990. 2. Apkon, M., and J. M. Nerbonne. Characterization of two distinct depolarization-activated K1 currents in isolated adult rat ventricular myocytes. J. Gen. Physiol. 97: 973–1010, 1991. 3. Capasso, J. M., A. Malhotra, R. M. Remily, J. Scheuer, and E. H. Sonnenblick. Effects of age on mechanical and electrical performance of rat myocardium. Am. J. Physiol. 245 (Heart Circ. Physiol. 14): H72–H81, 1983. 4. Dillmann, W. H. Diabetes mellitus induces changes in cardiac myosin in the rat. Diabetes 29: 579–582, 1980. 5. Dixon, J. E., W. Shi, H.-S. Wang, C. MacDonald, H. Yu, R. S. Wymore, I. S. Cohen, and D. McKinnon. Role of the Kv 4.3 K1 channel in ventricular muscle: a molecular correlate for the transient outward current. Circ. Res. 79: 659–668, 1996. Downloaded from http://ajpendo.physiology.org/ by 10.220.33.1 on May 2, 2017 shifted several mV negatively, and significantly, compared with 17- to 19-mo normal rat myocytes, without any change in k. The time course of recovery from inactivation of Ito was also compared between the diabetic and agematched control rat myocytes by use of a double-pulse method. Double pulses to 170 mV from a holding potential of 270 mV (each pulse duration 300 ms) were applied every 10 s, while the interpulse interval was increased from 10 ms to 1 s. The relative amplitude of Ito induced by the second pulse vs. the Ito amplitude induced by the first pulse was plotted against the interpulse interval. Recovery from its inactivation was not different between diabetic myocytes and agematched control rat myocytes, as shown in Fig. 4. In addition, the recovery of Ito was not altered in normal 17- to 19-mo rat myocytes compared with 2-mo rat myocytes. The previous studies showed that the kinetics of steady-state inactivation and recovery kinetics from inactivation were not altered in STZ-treated rats with short treatment periods of 1.5–2 mo (12) but were slowed in long-term (5- to 7-mo) STZ-treated rats (28). It seems that the kinetic changes of Ito may occur in the severely diseased myocardium with long-term STZinduced diabetes. Effects of diabetes on inward rectifying current. The current amplitudes, elicited by hyper- and depolarizing steps from 270 mV to up to 210 to about 2110 mV, were not different between diabetic rat myocytes and age-matched control rat myocytes (Fig. 5). Furthermore, the terminal component of the outward current elicited by depolarizing test pulses up to ,0 mV from a holding potential of 270 mV was not altered in diabetic rat myocytes compared with age-matched control myocytes (Fig. 2B). These results suggest that the inward rectifying potassium current (IK1 ) was not altered in the diabetic state. No alteration in the IK1 has also been demonstrated by other authors in STZ-treated diabetic rats (12, 17, 20), whereas reduced IK1 was reported in other diseased cardiac myocytes (13). Summary of results and significance of the decrease in Ito. The present study has, first of all, demonstrated that the current density of Ito in ventricular myocytes isolated from genetically diabetic rats sharing the characteristics with NIDDM was significantly less than that from 17- to 19-mo-old matched control rats. The current density of Ito from aged (17- to 19-mo) rats was significantly less than that from young adult (2-mo) rats. Thus the Ito density was reduced by aging, and the reduction was further accelerated by a diabetic state. Ito is considered to be one of the most important repolarizing currents in rat myocytes (11). The present study indicates that the lower Ito density, but not kinetic changes of the Ito, may be responsible for the longer duration of action potential in WBN/Kob rats, as already shown in our previous study (25). Shimoni et al. (24) demonstrated that the diabetic state exerted differential effects on the Ito in epicardial and endocardial myocytes from the left ventricle of short-term (6- to 7-day) STZ-treated rats. Such regional differences were not a focus of the present study but may be important in E699 E700 POTASSIUM CURRENTS IN GENETICALLY DIABETIC HEARTS 18. Makino, N., K. S. Dhalla, V. Elimban, and N. S. Dhalla. Sarcolemmal Ca21 transport in streptozotocin-induced diabetic cardiomyopathy in rats. Am. J. Physiol. 253 (Endocrinol. Metab. 16): E202–E207, 1987. 19. Nakama, K., K. Shichinohe, K. Kobayashi, K. Naito, O., Uchida, K. Yasuhara, and M. Tobe. Spontaneous diabetes-like syndrome in WBN/Kob rats. Acta Diabet. Lat. 22: 335–342, 1985. 20. Nobe, S., M. Aomine, M. Arita, S. Ito, and R. Takaki. Chronic diabetes mellitus prolongs action potential duration of rat ventricular muscles: circumstantial evidence for impaired Ca21 channel. Cardiovasc. Res. 24: 381–389, 1990. 21. Noda, N., H. Hayashi, H. Miyata, S. Suzuki, A. Kobayashi, and N. Yamazaki. Cytosolic Ca21 concentration and pH of rat myocytes during metabolic inhibition. J. Mol. Cell. Cardiol. 24: 435–445, 1992. 22. Pierce, G. N., J. B. Kutryk, and N. S. Dhalla. Alterations in Ca21 binding by and composition of the cardiac sarcolemmal membrane in chronic diabetes. Proc. Natl. Acad. Sci. USA 80: 5412–5416, 1983. 23. Schaffer, S. W., B. H. Tan, and G. L. Wilson. Development of a cardiomyopathy in a model of noninsulin-dependent diabetes. Am. J. Physiol. 248 (Heart Circ. Physiol. 17): H179–H185, 1985. 24. Shimoni, Y., D. Severson, and W. Giles. Thyroid status and diabetes modulate regional differences in potassium currents in rat ventricle. J. Physiol. Lond. 488: 673–688, 1995. 25. Tsuchida, K., H. Watajima, and S. Otomo. Calcium current in rat diabetic ventricular myocytes. Am. J. Physiol. 267 (Heart Circ. Physiol. 36): H2280–H2289, 1994. 26. Tsuchitani, M., T. Saegusa, I. Narama, T. Nishikawa, and T. Gonda. A new diabetic strain of rat (WBN/Kob). Lab. Anim. 19: 200–207, 1985. 27. Walker, K. E., E. G. Lakatta, and S. R. Houser. Age associated changes in membrane currents in rat ventricular myocytes. Cardiovasc. Res. 27: 1968–1977, 1993. 28. Wang, D. W., T. Kiyosue, S. Shigematsu, and M. Arita. Abnormalities of K1 and Ca21 currents in ventricular myocytes from rats with chronic diabetes. Am. J. Physiol. 269 (Heart Circ. Physiol. 38): H1288–H1296, 1995. 29. Wei, J. Y., H. A. Spurgeon, and E. G. Lakatta. Excitationcontraction in rat myocardium: alterations with adult aging. Am. J. Physiol. 246 (Heart Circ. Physiol. 15): H784–H791, 1984. 30. Xu, Z., K. P. Patel, and G. J. Rozanski. Metabolic basis of decreased transient outward K1 current in ventricular myocytes from diabetic rats. Am. J. Physiol. 271 (Heart Circ. Physiol. 40): H2190–H2196, 1996. Downloaded from http://ajpendo.physiology.org/ by 10.220.33.1 on May 2, 2017 6. Fein, F. S., I. B. Kornstein., J. E. Strobeck., J. M. Capasso, and E. H. Sonnenblick. Altered myocardial mechanics in diabetic rats. Circ. Res. 47: 922–933, 1980. 7. Gøtzsche, O. Myocardial cell dysfunction in diabetics mellitus. A review of clinical and experimental studies. Diabetes 35: 1158–1162, 1986. 8. Hamill, O. P., A. Marty, E. Neher, B. Sakmann, and F. J. Sigworth. Improved patch-clamp technique for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch. 391: 85–100, 1981. 9. Heyliger, C. E., A. Prakash, and J. H. McNeill. Alterations in cardiac sarcolemmal Ca21 pump activity during diabetes mellitus. Am. J. Physiol. 252 (Heart Circ. Physiol. 21): H540–H544, 1987. 10. Isenberg, G., U. Klockner, D. Mascher, and, U. Ravens. Changes in contractility and membrane currents as studied with a single patch-electrode whole-cell clamp technique. In: Electrophysiology of Single Cardiac Cells, edited by D. Noble and T. Powell. London: Academic, 1987, p. 26–67. 11. Josephson, I. R., J. Sanchez-Chapula, and A. M. Brown. Early outward current in rat single ventricular cells. Circ. Res. 54: 157–162, 1984. 12. Jourdon, P., and D. Feuvray. Calcium and potassium currents in ventricular myocytes isolated from diabetic rats. J. Physiol. Lond. 470: 411–429, 1993. 13. Kääb, S., B. Nuss, N. Chiamvimonvat, B. O’Rourke, P. H. Pak, D. A. Kass, E. Marban, and G. F. Tomaselli. Ionic mechanism of action potential prolongation in ventricular myocytes from dogs with pacing-induced heart failure. Circ. Res. 78: 262–273, 1996. 14. Ku, D. D., and B. M. Sellers. Effects of streptozotocin diabetes and insulin treatment on myocardial sodium pump and contractility of the rat heart. J. Pharmacol. Exp. Ther. 222: 395–400, 1982. 15. Lagadic-Gossmann, D., J. M. Chesnais, and D. Feuvray. Intracellular pH regulation in papillary muscle cells from streptozotocin diabetic rats: an ion sensitive microelectrode study. Pflügers Arch. 412: 613–617, 1988. 16. Legaye, F., P. Biegelman, E. Deroubaix, and E. Coraboeuf. Effect of 3,5,38-triiodothyronine treatment on cardiac action potential of streptozotocin-induced diabetic rat. Life Sci. 42: 2269–2274, 1988. 17. Magyar, J., Z. Rusznak, P. Szentesi, G. Szucs, and L. Kovacs. Action potentials and potassium currents in rat ventricular muscle during experimental diabetes. J. Mol. Cell. Cardiol. 24: 841–853, 1992.