Cellular mechanism of contractile dysfunction in the diabetic heart

Abstract

The aim of this study was to investigate the cellular mechanism(s) that underpins contractile dysfunction in the streptozotocin (STZ)-induced diabetic rat heart compared to age-matched control heart. In some experiments, a clinically relevant concentration of the volatile anaesthetic halothane (0.6 mM) was used examine its effect on contractile properties of STZ-induced diabetic heart. Diabetes was induced in male Wistar rats by a single i.p. injection of STZ (60 mg Kg-1, body weight) which, resulted in an experimental model of type 1 diabetes that was characterised by hypoinsulinaemia, hyperglycaemia, increases in osmolarity and decreases in body and heart weights. Total cation contents (Ca2+ Cu2+ Zn2+ Fe2+) were significantly (P<0.05) increased in the STZ-induced diabetic heart compared to age-matched controls. The majority of experiments were carried out on ventricular myocytes following 8-12 weeks of STZ treatment. L-type calcium (Ca2+) current (ICa,L) was measured in patch clamped ventricular myocytes in whole cell mode, using a cesium-based pipette solution and a holding potential of -40 mV and test potentials between —30 and 50 mV. The amplitude of ICal. was significantly (P<0.05) decreased in the STZ-induced diabetic myocytes compared to age-matched control. Furthermore, halothane further reduced the peak ICal, to levels in both age-matched control STZ-induced diabetic myocytes. Contraction was measured in electrically stimulated myocytes via a video-edge detector and results showed that the amplitude of contraction as a percentage of resting cell length (% RCL) was significantly (P<0.01) greater in STZ-induced diabetic myocytes (6.8 ± 0.5 %, n32) compared to that of age matched control (4.1 ± 1.04 %, n=27). Moreover, the 1pk of contraction was found to be significantly (P<0.01) longer in diabetic myocytes (164.1 ± 7.4 ms, n=30 Vs. 132.3 ± 5.9 ms, n=27) compared to control, respectively. Halothane evoked significant (P<0.05) reductions in the amplitude of contraction in control myocytes. The amplitude of contraction was significantly (P<0.01) reduced further in STZ-induced myocytes compared to the response in the absence of halothane. In voltage clamped myocytes however, contraction was peak amplitude of contraction was greater in control compared to STZ induced myocytes. Since contraction is ultimately dependent on cytosolic Ca 2+, it was relevant to measured free intracellular Ca 2+ concentrations ([Ca 2+]) using the fluorescent dye fura-2. Basal resting Ca2+ (measure by fluorescence ratio units) was significantly (P<0.01) increased in STZ-induced diabetic myocytes following 8 weeks of treatment compared to age-matched control (0.599 ± 0.009 ratio units, n=23 Vs. 0.521 ± 0.012 ratio units, n=23) , respectively. Electrically stimulated cardiac myocytes (1 Hz) induced Ca 2+ transients that had a longer time from the peak (tpk) of Ca2+ transient to half decay ( 1 decrn). Moreover, in the presence of halothane, the amplitude of electrically stimulated Ca 2+ transient release was significantly (P<0.05) decreased in control and STZ-induced myocytes but was not significantly altered between control and STZ-induced myocytes. Following a caffeine-induced Ca 2+ release, 1 1/2 of Ca decay was significantly (P<0.01) longer (43%) in myocytesobtained from STZ-induced compared to age-matched controls. However, in the presence of 10 mM nickel chloride (NiCl 2), the rate of Ca 2+ efflux out of the cell was similar in both control and diabetic myocyte. Myofilament sensitivity was studied by plotting the relationship between contraction and Ca 2+ in controland STZ-induced diabetic myocytes. The results show that myofilament sensitivity for Ca2+ is increased in the STZ-induced myocytes but is significantly (P<0.05) reduced following the application of halothane.

In conclusion, the results have shown that in electrically stimulated STZ-induced diabetic myocytes, the increase in contraction is primarily caused by an increase in myofilament Ca2+ sensitivity, and not through an increase in Ca 2+ release from the SR. Moreover, in the STZ-induced diabetic myocytes an alteration in Na+/Ca2+ -exchanger may contribute to a prolonged Ca2+ transient. It is suggested that prolonged Ap duration in the diabetic heart leads to increased Ca 2+ influx albeit a reduced lCa.t. which may overcompensate for a decrease in SERCA function (that has been reported in the diabetic heart. Misra el at 1999) and may lead to similar SR Ca 2 load and release in both diabetic and control myocytes. Following. SR Ca 2+ release it is suggested that the increased myofilament Ca 2+ sensitivity in STZ-induced myocytes leads to an increase in contraction that has been reported in this study.

In voltage clamped STZ-induced diabetic myocytes, a decrease in lCa.l. was mirrored by a decrease in the peak amplitude of contraction. It is suggested that in voltage clamped myocytes from STZ-induced hearts, that are not influenced by the Ap, decreased lCa.L, may lead to a reduced Ca 2+ influx and subsequent SR Ca2+ release. Reduced Ca 2+ release from the SR, may not be compensated for by the increase in myofilament Ca 2+ sensitivity in the diabetic heart, and may ultimately lead to a reduction in the amplitude of contraction that has been reported in this study.

It has also been shown that, following the application of halothane. the lCa.L, Ca2+ transient and amplitude of contraction were significantly more decreased in STZ-induced myocytes compared to that of control. It is suggested that reduced myofilament Ca2+ sensitivity in the presence of halothane contributes to the changes in contraction. However, it is also likely that another mechanism such as fractional Ca 2+ release and/or SR Ca2+ load may also be affected by the actions of halothane in the diabetic heart.