Table of Contents  

Howarth, Qureshi, Hassan, Al Kury, Isaev, Parekh, Yammahi, John, Oz, Raza, Adeghate, and Adrian: Ventricular myocyte contraction, intracellular calcium and expression of genes encoding cardiac muscle proteins in young and ageing Zucker diabetic fatty rat heart

Introduction

Diabetes mellitus and its complications are a public health problem of epidemic proportions. The total number of people with diabetes is projected to rise from 171 million in 2000 to 366 million in 2030.1 The association between type 2 diabetes mellitus and obesity is very strong and cardiovascular disease is the leading cause of morbidity and mortality among diabetic patients.2,3 A recent study among Emirati citizens reported that the age-standardised rates for diabetes mellitus (diagnosed and undiagnosed) and pre-diabetes among those 30–64 years old were 29.0% and 24.2%, respectively.4

Doppler imaging, echocardiography, radionuclide angiography and other techniques have demonstrated a variety of diastolic and systolic dysfunctions in type 2 diabetic patients. Haemodynamic abnormalities include reduced left ventricular ejection fraction, impaired myocardial velocity at early diastole, abnormal relaxation during the early filling phase, prolonged isovolumetric relaxation, lower peak systolic and early diastolic velocity, impaired diastolic relaxation and filling and reduced peak filling rate, with the severity of the abnormalities depending on the patients’ age and duration of diabetes.5

The Zucker diabetic fatty (ZDF) rat is a genetic model in which the male homozygous (FA/FA) animals develop obesity and type 2 diabetes mellitus.6 As in obese humans, ZDF rats exhibit early β-cell compensation (hyperplasia) of insulin resistance followed by decompensation (loss of cells).7 The early changes in β-cell responsiveness to glucose may contribute to the hyperinsulinaemia and subsequent insulin resistance.8 Heart function is compromised in the ZDF rat. Recent studies performed in young (9–13 weeks) and ageing (30–34 weeks) ZDF rats have variously demonstrated a prolonged time course of myocyte shortening and relaxation which was associated with prolonged time course of the Ca2+ transient, a reduced amplitude of L-type Ca2+ current, and changed expression of genes encoding a variety of L-type Ca2+ channel, cardiac muscle, intracellular Ca2+ transport and regulatory and cardiac muscle proteins.9,10 The changes in ventricular myocyte contraction, intracellular calcium and the expression of genes encoding cardiac muscle proteins that take place in young (9–13 weeks) and ageing (30–34 weeks) Zucker diabetic fatty rat heart are reviewed here.

Materials and methods

Ventricular myocyte isolation

Experiments were performed in young (9–13 weeks) and ageing (30–34 weeks) male ZDF (FA/FA) rats and age-matched Zucker lean (ZL; +/FA) controls (Charles River Laboratories, Margate, Kent, UK). Approval for the project was obtained from the Animal Ethics Committee, Faculty of Medicine and Health Sciences, United Arab Emirates University, United Arab Emirates. Left ventricular myocytes were isolated according to previously described techniques.9,10 Animals were euthanized using a guillotine. Hearts were removed rapidly, mounted on a Langendorff system and then perfused at a constant flow of 8 ml per gram of heart tissue per minute and at 36–37°C with a cell isolation solution containing (in mmol/l): 130.0 NaCl, 5.4 KCl, 1.4 MgCl2, 0.75 CaCl2, 0.4 NaH2PO4, 5.0 HEPES, 10.0 glucose, 20.0 taurine and 10.0 creatine (pH 7.3). The perfusion flow rate was adjusted to allow for differences in heart weight between animals. When the heart had stabilized, perfusion was continued for 4 min with Ca2+-free cell isolation solution containing 0.1 mmol/l EGTA and then for 6 min with cell isolation solution containing 0.05 mmol/l Ca2+, 0.75 mg/ml type 1 collagenase (Worthington Biochemical Corp., Lakewood, NJ, USA) and 0.075 mg/ml type XIV protease (Sigma, Taufkirchen, Germany). Left ventricle tissue was excised from the heart, minced and gently shaken in collagenase-containing isolation solution supplemented with 1% bovine serum albumin (BSA). Cells were filtered from this solution at 4-min intervals and resuspended in cell isolation solution containing 0.75 mmol/l Ca2+.

Measurement of ventricular myocyte shortening

Ventricular myocytes were allowed to settle on the glass bottom of a Perspex chamber mounted on the stage of an inverted Axiovert 35 microscope (Zeiss, Göttingen, Germany). Cells were superfused (3–5 ml/min) with normal Tyrode containing the following (in mmol/l): 140.0 NaCl, 5.0 KCl, 1.0 MgCl2, 10.0 glucose, 5.0 HEPES and 1.8 CaCl2 (pH 7.4). Unloaded myocyte shortening was recorded using a VED-114 video-edge detection system (Crystal Biotech, Northborough, MA, USA). Resting cell length, time to peak (TPK) shortening, time from peak to half (THALF) relaxation and amplitude of shortening (expressed as a percentage of resting cell length) were measured in field-stimulated (1 Hz) myocytes maintained at 35–36°C. Data were acquired and analysed with Signal Averager software version 6.37 (Cambridge Electronic Design, Cambridge, UK).

Measurement of intracellular Ca2+ concentration

Myocytes were loaded with the fluorescent indicator fura-2 AM (F-1221, Molecular Probes, Eugene, OR, USA) as described previously.9,10 To measure intracellular Ca2+ concentration, myocytes were alternately illuminated by 340-nm and 380-nm light using a monochromator (Cairn Research, Faversham, UK) which changed the excitation light every 2 msec. The resulting fluorescence, emitted at 510 nm, was recorded by a photomultiplier tube and the ratio of the emitted fluorescence at the two excitation wavelengths (340 : 380) was calculated to provide an index of intracellular Ca2+ concentration. The resting fura-2 ratio, TPK Ca2+ transient, THALF decay of the Ca2+ transient and the amplitude of the Ca2+ transient were measured in field-stimulated (1 Hz) myocytes maintained at 35–36°C.

Measurement of L-type Ca2+ current

Voltage-dependent L-type Ca2+ current was measured using previously described whole-cell patch-clamp techniques.9,10 L-type Ca2+ current was recorded with an Axopatch 200B amplifier (Molecular Devices, Sunnyvale, CA, USA). The analogue signal was filtered using an eight-pole Bessel filter with a bandwidth of 5 kHz and digitized at a sampling rate of 50 kHz. Patch pipettes were fabricated from filamented GC150TF borosilicate glass (Harvard Apparatus, Holliston, MA, USA). Electrode resistances ranged from 1 to 1.8 MΩ, and seal resistances were 1–5 GΩ. Whole-cell voltage-clamp data were elicited from a holding potential of −40 mV to membrane potentials ranging from −30 to +70 mV. Inactivation of current was measured as the relation of amplitude of the current at 100 msec after peak current to the amplitude of the peak current at different test potentials. The whole-cell bath solution contained the following (in mmol/l): 95 NaCl, 50 TEA Cl, 2.0 MgCl2, 2.0 CaCl2, 10.0 HEPES and 10 glucose (pH 7.35). The pipette solution contained the following (in mmol/l): 140 CsCl, 2.0 MgCl2, 1.0 MgATP, 10.0 EGTA, 10.0 TRIS (pH 7.25). Experiments were performed at room temperature (22–24°C). Data were analysed using computer-fitting software Origin™ version 7.1 (Microcal Software, Northampton, MA, USA).

Measurement of sarcoplasmic reticulum Ca2+ transport

Sarcoplasmic reticulum (SR) Ca2+ release was assessed using previously described techniques.9,10 After establishing steady state Ca2+ transients in field-stimulated (1 Hz) myocytes maintained at 35–36°C and loaded with fura-2, stimulation was paused for a period of 5 sec. Caffeine (20 mM) was then applied for 10 sec using a solution-switching device customized for rapid solution exchange.11 Electrical stimulation was then resumed and the Ca2+ transients were allowed to recover to steady state. Fractional release of SR Ca2+ was assessed by comparing the amplitude of the electrically evoked steady-state Ca2+ transients with that of the caffeine-evoked Ca2+ transient. Ca2+ refilling of SR was assessed by measuring the rate of recovery of electrically evoked Ca2+ transients following application of caffeine.

Assessment of myofilament sensitivity to Ca2+

In some cells shortening and fura-2 ratio were recorded simultaneously. Myofilament sensitivity to Ca2+ was assessed from phase–plane diagrams of fura-2 ratio compared with cell length by measuring the gradient of the fura-2 –cell-length trajectory during late relaxation of the twitch contraction as previously described.9,10 The position of the trajectory reflects the relative myofilament response to Ca2+ and hence can be used as a measure of myofilament sensitivity to Ca2+.12

Gene expression

Expression of genes encoding a variety of muscle proteins was evaluated using previously described techniques.9,10 Samples of left ventricle tissue were collected from the apex of the heart in vivo and immediately placed in RNAlater (AM7021; Applied Biosystems, Carlsbad, CA, USA), kept for 24 hours at room temperature and then frozen at −80°C pending further processing. Total RNA was extracted from homogenized tissue using an RNeasy mini kit (74104, Qiagen, Valencia, CA, USA). The concentration and purity of the RNA samples were determined by measuring the absorbance at 260 nm (A260) and the ratio of the absorbance at 260 nm and 280 nm (ND-1000 NanoDrop). Total RNA was converted into complementary DNA (cDNA) using a high capacity cDNA reverse transcription (RT) kit (4374966; Applied Biosystems, Carlsbad, CA, USA). Gene expression assays were performed using custom Taqman low-density arrays (32 Format 4346799; Applied Biosystems, Carlsbad, CA, USA). Each assay on the low-density-array panel was spotted in triplicate for each RNA sample. The target genes for real-time RT–polymerase chain reaction (RT-PCR) analyses are listed in Table 1. Rat hypoxanthine guanine phosphoribosyl transferase was used as an endogenous control. One hundred nanograms of cDNA (RNA equivalent) were loaded together with 2 × TaqMan gene expression master mix (No AmpErase UNG; Applied Biosystems, Carlsbad, CA, USA) for a total of 100 μl per port. Real-time RT-PCR was performed in a 7900HT Fast Real-time PCR System (Applied Biosystems, Carlsbad, CA, USA).

TABLE 1

Target genes and alternative names of genes selected for real-time RT-PCR analyses

Cardiac muscle proteins and associated regulatory proteins
MYH6 Myosin, heavy polypeptide 6, cardiac muscle, alpha
MYH7 Myosin, heavy polypeptide 7, cardiac muscle, beta
MYL2 Myosin, light polypeptide 2, regulatory, cardiac, slow
MYBPC3 Myosin binding protein C, cardiac
ACTC1 Actin, alpha, cardiac muscle 1
TNNI3 Troponin I type 3, cardiac
TNNT2 Troponin T2, cardiac
TNNC1 Troponin C, cardiac/slow skeletal
TNNC2 Troponin C2, fast
Cell membrane ion transport
ATP1A1 ATPase, Na+/K+ transporting, alpha 1 polypeptide
ATP1B1 ATPase, Na+/K+ transporting, beta 1 polypeptide
SLC8A1 Solute carrier family 8 (Na+–Ca2+ exchanger)
SLC9A1 Solute carrier family 9 (Na+–H+ exchanger), member 1
CACNA1C Calcium channel, voltage-dependent, L type, alpha 1C subunit
CACNA1D Calcium channel, voltage-dependent, L type, alpha 1D subunit
CACNA2D1 Calcium channel, voltage-dependent, alpha2/delta subunit 1
CACNA2D2 Calcium channel, voltage-dependent, alpha 2/delta subunit 2
CACNA2D3 Calcium channel, voltage-dependent, alpha2/delta subunit 3
CACNB2 Calcium channel, voltage-dependent, beta 2 subunit
Intracellular Ca2+ transport and Ca2+ regulation
ATP2A2 ATPase, Ca2+-transporting, cardiac muscle, slow twitch 2
ATP2A1 ATPase, Ca2+-transporting, cardiac muscle, fast twitch 1
RYR2 Ryanodine receptor 2, cardiac
CASQ2 Calsequestrin 2
PLN Phospholamban
CALM1 Calmodulin 1
CALM2 Calmodulin 2
PRKAA2 Protein kinase, AMP-activated, alpha 2 catalytic subunit

Statistics

Results were expressed as the mean ± standard error of the mean (S.E.M.). Statistical comparisons were performed using either the independent samples t-test or one-way ANOVA followed by Bonferroni-corrected test for multiple comparisons, as appropriate. P ≤ 0.05 was considered to indicate a significant difference.

Results

General characteristics

Diabetes in young and ageing ZDF rats was characterized by a fourfold increase in non-fasting blood glucose compared with age-matched controls. Body weight was significantly (P < 0.01) higher in young ZDF and was slightly lower in ageing ZDF rats compared with controls. Heart weight was significantly (P < 0.01) higher in young ZDF and slightly lower in ageing ZDF compared with controls. Heart weight to body weight ratio was significantly (P < 0.01) lower in young and slightly higher in ageing ZDF rats compared with controls.

Ventricular myocyte shortening

Resting cell length was not significantly (P >0.01) altered in young and significantly (P <0.01) reduced in ageing ZDF myocytes compared with ZL controls. Time to peak shortening and THALF relaxation of shortening were significantly (P <0.01) prolonged in young and not significantly (P >0.05) altered in ageing ZDF myocytes. In young animals, TPK was 163 ± 5 msec in ZDF myocytes compared with 136 ± 5 msec in controls9. In ageing animals, TPK was 141 ± 5 msec in ZDF myocytes compared with 132 ± 3 msec in controls.10 In young animals, THALF relaxation of shortening was 127 ± 7 msec in ZDF myocytes compared with 103 ± 4 msec in controls9. In ageing animals, THALF relaxation of shortening was 85 ± 6 msec in ZDF myocytes compared with 86 ± 4 msec in controls.10 Although the amplitude of shortening was slightly increased in young ZDF and slightly reduced in ageing ZDF myocytes, the difference compared with controls was not significant (P > 0.05) (Figure 1).

FIGURE 1

Ventricular myocyte shortening. Mean amplitude of shortening (expressed as a percentage of control) in myocytes from young and ageing ZDF rats compared with age-matched controls. Experiments were performed in electrically stimulated myocytes maintained at 35–36°C. Data are mean, n = 56–70 cells from 6–8 hearts.

5-2-4-fig1.jpg

Intracellular Ca2+

Resting fura-2 ratio was significantly (P < 0.05) elevated in young ZDF myocytes and not significantly (P > 0.05) altered in ageing ZDF myocytes compared with controls. In young and ageing animals, TPK Ca2+ transient was significantly (P < 0.01) prolonged in ZDF myocytes compared with controls. In young animals, TPK was 67 ± 3 msec compared with 58 ± 2 msec in ZL controls.9 In ageing animals, TPK was 70 ± 3 msec in ZDF myocytes compared with 58 ± 2 msec in ZL controls.10 In young and ageing animals, THALF decay of the Ca2+ transient was not significantly (P > 0.05) altered in ZDF myocytes compared with controls. Although the amplitude of the Ca2+ transient was slightly increased in young ZDF and slightly reduced in ageing ZDF myocytes, the difference compared with age-matched controls was not significant (P > 0.05) (Figure 2).

FIGURE 2

Ventricular myocyte intracellular Ca2+. Mean amplitude of the Ca2+ transient (expressed as a percentage of control) in myocytes from young and ageing ZDF rats compared with age-matched controls. Experiments were performed in electrically stimulated myocytes maintained at 35–36°C. Data are mean, n = 48–55 cells from 7–9 hearts.

5-2-4-fig2.jpg

L-type Ca2+ current

The amplitude of L-type Ca2+ current was significantly (P <0.05) reduced in myocytes from young and ageing ZDF rats compared with controls (Figure 3). In young animals, at a test potential of 0 mV, the density of L-type Ca2+ current was 1.2 ± 0.3 pA pF−1 in ZDF myocytes compared with 2.4 ± 0.4 pA pF−1 in controls.9 In ageing animals, the density of L-type Ca2+ current was 0.8 ± 0.1 pA pF–1 in ZDF myocytes compared with 2.2 ± 0.3 pA pF−1 in controls.10

FIGURE 3

L-type Ca2+ current. Mean amplitude of L-type Ca2+ current (expressed as a percentage of control) in myocytes from young and ageing ZDF rats compared with age-matched ZL controls during a test pulse to 0 mV from a holding potential of −40 mV. Experiments were performed in myocytes maintained at 22–24°C. Data are mean, n = 5–18 cells from six hearts.

5-2-4-fig3.jpg

Sarcoplasmic reticulum Ca2+ transport and myofilament sensitivity to Ca2+

Amplitude of caffeine-evoked Ca2+ release and SR Ca2+ fractional release (amplitude of electrically evoked compared with caffeine-evoked Ca2+ transient) was not significantly (P > 0.05) altered in myocytes from young and ageing ZDF rats compared with agematched ZL controls. Recovery of the Ca2+ transient, after application of caffeine and following resumption of electrical stimulation was not significantly (P > 0.05) altered in myocytes from young and ageing ZDF animals compared with controls.

The gradient of the fura-2 cell-length trajectory during late relaxation (500–800 msec) of the twitch contraction, a measure of myofilament sensitivity to Ca2+, was not significantly (P > 0.05) altered during the periods 500–600, 500–700 and 500–800 msec in myocytes from young ZDF animals and 500–800 and 500–600 msec in myocytes from ageing ZDF animals compared with controls. However, there was a significant (P < 0.05) alteration in gradient during the period 500–700 msec in myocytes from ageing ZDF animals compared with controls.9,10

Expression of genes encoding Ca2+ transport and muscle proteins

Expression of some genes encoding Ca2+ channels, plasma membrane transporters, SR Ca2+ regulatory and cardiac muscle proteins were variously up-regulated, down-regulated or unchanged in young and ageing ZDF ventricle compared with age-matched ZL controls. The changes are shown in Table 2.9,10

TABLE 2

Changes in gene expression of Ca2+ channels, plasma membrane transporters, SR Ca2+ regulatory and cardiac muscle proteins in young and ageing ZDF rat heart compared with controls

Target gene Young ZDF Ageing ZDF
Ca2+ channels
CANNA1C Increased**
CACNA1D
CACNA1G Increased* Increased*
CACNA1H Increased* Increased**
CACNA2D1 Increased*
CACNA2D2
CACNA2D3 Decreased**
Plasma membrane transporters
ATP1A1 Increased**
ATP1B1 Decreased**
SLC8A1
SLC9A1 Decreased** Decreased*
SR Ca2+ and regulatory proteins
ATP2A1 Increased**
ATP2A2 Decreased** Decreased**
RYR2
CASQ2
PLN
CALM1 Decreased**
CALM2
PRKAA2
Cardiac muscle and regulatory proteins
MYH6 Decreased** Decreased**
MYH7 Increased**
MYL2 Decreased* Increased*
MYBPC3
ACTC1 Decreased*
TNNI3 Decreased**
TNNT2 Decreased** Decreased**
TNNC1 Decreased**
TNNC2

*, P < 0.05; **, P < 0.01.

Discussion

Non-fasting blood glucose was elevated fourfold in young and ageing ZDF rats compared with age-matched ZL controls. Previous studies have demonstrated that in addition to hyperglycaemia the ZDF metabolic phenotype also displays raised plasma haemoglobin A1c, impaired glucose tolerance, hyperinsulinaemia, dyslipidaemia, reduced myocardial glucose uptake and reduced rates of glucose oxidation1318. Cardiac fatty acid utilization is enhanced in ZDF rat hearts with increased rates of fatty acid oxidation and elevated levels of myocardial triacylglycerols. Enhanced cardiac fatty acid utilization in rodent models of type 2 diabetes mellitus is associated with reduced cardiac contractile function, perhaps as a consequence of lipotoxicity and/or reduced cardiac efficiency.19 Body weight and heart weight were significantly increased in young ZDF rats compared with age-matched controls. Although the resting cell length (RCL) of myocytes from young ZDF rats was unaltered, previous studies have demonstrated an increase in cell volume accompanied by increased left ventricular atrial natriuretic peptide (ANP) during the early stages of diabetic cardiomyopathy.20 In ageing ZDF, rat body weight and heart weight was a little smaller and RCL of myocytes was significantly smaller compared with age-matched controls. The reduced body weight might partly be associated with the ongoing metabolic disturbance of carbohydrates metabolism and an increasing utilization of fatty acid metabolism to meet energy requirements this might in turn lead to a progressive drain in fat reserves and loss of body weight.

Time to peak shortening and THALF relaxation of shortening were prolonged in myocytes from young and not-significantly altered in ageing ZDF animals compared with age-matched controls. Amplitude of shortening was slightly increased in myocytes from young and slightly reduced in myocytes from ageing ZDF animals compared with controls; however, these small changes were not significant. Previous studies in animals aged 22 weeks demonstrated impaired relaxation in myocytes from ZDF animals compared with age-matched lean controls21 and, in animals aged 30 weeks, reduced left ventricular contraction, relaxation and developed pressure in hearts from ZDF rats compared with age-matched controls.22 Echocardiography has demonstrated diastolic and systolic dysfunction which was paralleled by decreased maximal force and maximal rate of force redevelopment in myocytes in ZDF rats compared with controls. The myocardial functional changes were associated with whole-body insulin sensitivity, decreased myocardial glucose utilization, decreased glucose transporter-4 mRNA and protein expression and an increased abundance of the fatty acid transporter CD36 in ZDF hearts compared with ZL controls.16 Lower myocardial glucose uptake has also been demonstrated at both week 14 and week 19 of age and this was associated with diminished expression of glucose transporter 4 (GLUT4) in ZDF rats compared with controls.17 Collectively, these results suggest that early alterations in myocardial substrate metabolism may contribute to myocardial dysfunction.

Cardiac muscle myosin is a hexamer consisting of two heavy-chain subunits, two light-chain subunits and two regulatory units. In young ZDF rats, expression of the gene encoding MYH6 (α-myosin) was down-regulated, whereas MYH7 (β-myosin) was up-regulated.9 However, in ageing ZDF rats, α-myosin was down-regulated, whereas β-myosin was unaltered.10 α-Myosin has a higher ATPase activity than β-myosin and the contractile velocity of the heart is correlated with the relative amount of each myosin heavy chain. Hearts expressing more α-myosin have more rapid cross-bridge cycling and a higher rate of contraction, whereas hearts with more β-myosin have a lower rate of contraction. Downregulation of α-myosin and up-regulation of β-myosin might partly underlie the prolonged time course of myocyte contraction in young ZDF rats.

In young ZDF rats, amplitude of myocyte shortening and amplitude of the Ca2+ transient were slightly increased, whereas in ageing ZDF rats amplitude of myocyte shortening was slightly reduced and amplitude of the Ca2+ transient was unaltered. The density of L-type Ca2+ current was significantly reduced over a wide range of test potentials in myocytes from young, and even more in myocytes from ageing, ZDF rats compared with controls. Changes in L-type Ca2+ current were associated with altered expression of genes encoding a variety of Ca2+ channel and regulatory proteins including CACNA1C, CACNA2D1 and CACNA2D3. If up-regulation of CACNA1C translated into up-regulation of the α1 subunit then this might alter the voltage sensitivity and, perhaps, the activation and/or inactivation properties of the L-type Ca2+ channel.

Despite the reductions in L-type Ca2+ current, SR Ca2+ transport and amplitude of shortening did not appear to be significantly altered in myocytes from young or ageing ZDF rats compared with controls. How might a reduced amplitude of L-type Ca2+ current be able to provide a trigger for what appears to be normal SR Ca2+ release and generation of a normal Ca2+ transient? One possibility might be increased sensitivity of the SR Ca2+ release channel. However, the expression of RYR2 was unaltered in young and ageing ZDF rat ventricles. Another possibility might be an influx of Ca2+ via the Na+/Ca2+ exchange, which together with influx of Ca2+ on the L-type Ca2+ channel may collectively be able to provide a normal trigger for SR Ca2+ release and, therefore, generation of a normal Ca2+ transient. Expression of the gene encoding Na+/Ca2+ exchange protein (SLC8A1) was unaltered in ventricles from young and ageing ZDF rats compared with age-matched controls. Further studies would be required to evaluate the electrophysiological function of the Na+/Ca2+ exchange in ZDF rat hearts.

A change of intracellular pH can have a profound effect on myocyte shortening and in particular myofilament sensitivity to Ca2+.23 Decreased Na+/H+ exchange activity has been previously demonstrated in diabetic rat hearts.24 Expression of the gene encoding Na+/H+ exchange (SLC9A1) was downregulated in both young and ageing ZDF rat hearts. However, with the exception of the 500–700 msec time point in ageing ZDF myocytes, there was little effect on myofilament sensitivity to Ca2+ in myocytes from either young or ageing ZDF rats compared with controls.

Conclusion

Age-dependent changes in myocyte contractility are accompanied by changes in Ca2+ transport, including a reduction in L-type Ca2+ current and subtle changes in expression of a variety of genes including the Ca2+ channel, membrane transporters, SR Ca2+ and cardiac muscle proteins.

Acknowledgements

This work was supported by a grant from Sheikh Hamdan Bin Rashid Al Maktoum Award for Medical Sciences.

References

1. 

Wild S, Roglic G, Green A, Sicree R, King H. Global prevalence of diabetes: estimates for the year 2000 and projections for 2030. Diabetes Care 2004; 27:1047–53. http://dx.doi.org/10.2337/diacare.27.5.1047

2. 

Zimmet PZ, Alberti KG. Introduction: Globalization and the non-communicable disease epidemic. Obesity (Silver Spring) 2006; 14:1–3.http://dx.doi.org/10.1038/oby.2006.1

3. 

Shoghi KI, Finck BN, Schechtman KB, Sharp T, Herrero P, Gropler RJ, et al. In vivo metabolic phenotyping of myocardial substrate metabolism in rodents: differential efficacy of metformin and rosiglitazone monotherapy. Circ Cardiovasc Imaging 2009; 2:373–81.http://dx.doi.org/10.1161/CIRCIMAGING.108.843227

4. 

Saadi H, Carruthers SG, Nagelkerke N, Al-Maskari F, Afandi B, Reed R, Lukic M, et al. Diabetes Res Clin Pract 2007; 78:369–77.http://dx.doi.org/10.1016/j.diabres.2007.04.008

5. 

von Bibra H, Thrainsdottir IS, Hansen A, Dounis V, Malmberg K, Ryden L. Tissue Doppler imaging for the detection and quantitation of myocardial dysfunction in patients with type 2 diabetes mellitus. Diab Vasc Dis Res 2005; 2:24–30.http://dx.doi.org/10.3132/dvdr.2005.002

6. 

Clark JB, Palmer CJ, Shaw WN. The diabetic Zucker fatty rat. Proc Soc Exp Biol Med 1983; 173(1):68–75.

7. 

Shimabukuro M, Zhou YT, Levi M, Unger RH. Fatty acid-induced beta cell apoptosis: a link between obesity and diabetes. Proc Natl Acad Sci USA 1998; 95:2498–502.http://dx.doi.org/10.1073/pnas.95.5.2498

8. 

Zawalich WS, Zawalich KC, Kelley GG, Shulman GI. Islet phosphoinositide hydrolysis and insulin secretory responses from prediabetic fa/fa ZDF rats. Biochem Biophys Res Commun 1995; 209:974–80.http://dx.doi.org/10.1006/bbrc.1995.1593

9. 

Howarth FC, Qureshi MA, Hassan Z, Al Kury LT, Isaev D, Parekh K, et al. Changing pattern of gene expression is associated with ventricular myocyte dysfunction and altered mechanisms of Ca2+ signalling in young type 2 Zucker diabetic fatty rat heart. Exp Physiol 2011; 96:325–37.http://dx.doi.org/10.1113/expphysiol.2010.055574

10. 

Howarth FC, Qureshi MA, Hassan Z, Isaev D, Parekh K, John A, et al. Contractility of ventricular myocytes is well preserved despite altered mechanisms of Ca(2+) transport and a changing pattern of mRNA in aged type 2 Zucker diabetic fatty rat heart. Mol Cell Biochem 2012; 361:267–80.http://dx.doi.org/10.1007/s11010-011-1112-y

11. 

Levi AJ, Hancox JC, Howarth FC, Croker J, Vinnicombe J. A method for making rapid changes of superfusate whilst maintaining temperature at 37 degrees C. Pflugers Arch 1996; 43:930–7.http://dx.doi.org/10.1007/s004240050217

12. 

Spurgeon HA, DuBell WH, Stern MD, Sollott SJ, Ziman BD, Silverman HS, et al. Cytosolic calcium and myofilaments in single rat cardiac myocytes achieve a dynamic equilibrium during twitch relaxation. J Physiol 1992; 447:83–102.

13. 

Welch MJ, Lewis JS, Kim J, Sharp TL, Dence CS, Gropler RJ, et al. Assessment of myocardial metabolism in diabetic rats using small-animal PET: a feasibility study. J Nucl Med 2006; 47:689–97.

14. 

Pontes Andersen CC, Holmstrup P, Buschard K, Flyvbjerg A. Renal alterations in prediabetic rats with periodontitis. J Periodontol 2008; 79:684–90.http://dx.doi.org/10.1902/jop.2008.070433

15. 

Forcheron F, Basset A, Del Carmine P, Beylot M. Lipase maturation factor 1: its expression in Zucker diabetic rats, and effects of metformin and fenofibrate. Diabetes Metab 2009; 35:452–7.http://dx.doi.org/10.1016/j.diabet.2009.05.004

16. 

van den Brom CE, Huisman MC, Vlasblom R, Boontje NM, Duijst S, Lubberink M, et al. Altered myocardial substrate metabolism is associated with myocardial dysfunction in early diabetic cardiomyopathy in rats: studies using positron emission tomography. Cardiovasc Diabetol 2009; 8:39.

17. 

Shoghi KI, Gropler RJ, Sharp T, Herrero P, Fettig N, Su Y, et al. Time course of alterations in myocardial glucose utilization in the Zucker diabetic fatty rat with correlation to gene expression of glucose transporters: a small-animal PET investigation. J Nucl Med 2008; 49:1320–1327.http://dx.doi.org/10.2967/jnumed.108.051672

18. 

Golfman LS, Wilson CR, Sharma S, Burgmaier M, Young ME, Guthrie PH, et al. Activation of PPARgamma enhances myocardial glucose oxidation and improves contractile function in isolated working hearts of ZDF rats. Am J Physiol Endocrinol Metab 2005; 289:E328–36.http://dx.doi.org/10.1152/ajpendo.00055.2005

19. 

Carley AN, Severson DL. Fatty acid metabolism is enhanced in type 2 diabetic hearts. Biochim Biophys Acta 2005; 1734:112–26.http://dx.doi.org/10.1016/j.bbalip.2005.03.005

20. 

Fredersdorf S, Thumann C, Ulucan C, Griese DP, Luchner A, Riegger GA, et al. Myocardial hypertrophy and enhanced left ventricular contractility in Zucker diabetic fatty rats. Cardiovasc Pathol 2004; 13:11–19.http://dx.doi.org/10.1016/S1054-8807(03)00109-1

21. 

Fulop N, Mason MM, Dutta K, Wang P, Davidoff AJ, Marchase RB, et al. Impact of Type 2 diabetes and ageing on cardiomyocyte function and O-linked N-acetylglucosamine levels in the heart. Am J Physiol Cell Physiol 2007; 292:C1370–8.http://dx.doi.org/10.1152/ajpcell.00422.2006

22. 

Burgdorf C, Richardt G, Schutte F, Dendorfer A, Kurz T. Impairment of presynaptic alpha2-adrenoceptor-regulated noradrenaline overflow in failing hearts from Zucker diabetic fatty rats. J Cardiovasc Pharmacol 2006; 47:256–62.http://dx.doi.org/10.1097/01.fjc.0000202560.61667.3e

23. 

Liou YM, Chang JC. Differential pH effect on calcium-induced conformational changes of cardiac troponin C complexed with cardiac and fast skeletal isoforms of troponin I and troponin T. J Biochem 2004; 136:683–92.http://dx.doi.org/10.1093/jb/mvh175

24. 

Feuvray D. Intracellular pH control mechanisms in the diabetic myocardium. Diabetes Res Clin Pract 1996; 31 (Suppl.):S87–92.





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