Sex differences in newborn myocardial metabolism and response to ischemia.

In children with congenital heart disease, female sex has been linked to greater in-hospital mortality associated with low cardiac output, yet the reasons for this are unclear. Therefore, we examined whether newborn sex differences in the heart's metabolic response to ischemia exist. Left ventricular (LV) in vivo and ischemic biopsies of newborn male and female piglets were compared. Tissue ATP, creatine phosphate (CP), glycogen, anaerobic end-products lactate and hydrogen ion (H), and key regulatory enzymes were measured. Compared with males, newborn females displayed 14% lower ATP, 22% lower CP, and 32% lower glycogen reserves (p < 0.05) at baseline. During ischemia, newborn females accumulated 17% greater lactate and 40% greater H accumulation (p < 0.02), which was associated with earlier cessation of glycolysis and lower ischemic ATP levels (p < 0.02) compared with males. Newborn females demonstrated a greater ability to use their glycogen reserves, resulting in significantly lower (p < 0.003) glycogen levels throughout the ischemic period. Thus, newborn females are at a metabolic disadvantage because they exhibited lower energy levels and greater tissue lactic acidosis, both linked to an increase susceptibility to ischemic injury and impair myocardial function on reperfusion.

Ventricle-specific metabolic differences in the newborn piglet myocardium in vivo and during arrested global ischemia.

Ventricular dysfunction is reported greater in the left (LV) versus right ventricle (RV) in infants following surgically induced ischemia. Ventricle-specific differences in baseline metabolism may alter response to ischemia thus affecting postischemic functional recovery. This study identifies ventricle-specific metabolic differences in the newborn (piglet) heart at baseline (working) and during ischemia (arrested). Baseline LV citrate synthase (CS) and hydroxyacyl-CoA dehydrogenase (HAD) activities were 15% and 18% lower (p < 0.02), whereas creatine kinase (CK) and phosphofructokinase (PFK) activities were 40% and 23% higher (p < 0.04) than the RV. Baseline LV glycogen reserves were also 55% higher (p = 0.004). By 15 min of ischemia, LV ATP was 20% lower (p < 0.05), lactate was 51% higher (p = 0.001), and hydrogen ions (H) were 43% higher (p = 0.03) compared with the RV. These differences persisted for the entire ischemic period (p < 0.02). After 45 min of ischemia, the LV used 58% less (p < 0.05) glycogen than the RV. These findings demonstrate that the enhanced glycolytic capacity of the newborn LV was accompanied by greater anaerobic end-product accumulation and lower energy levels during ischemia. This profile may offer one explanation for greater LV-dysfunction relative to the RV in children following ischemia.

Impact of female sex hormones on liver tissue lactic acidosis during ischemia.

BACKGROUND: Lower liver transplant success is observed when the donor is female. Intracellular acidosis during ischemia is proposed to contribute to the injury sustained by the transplanted organ and its role in livers obtained from nonheartbeating donors is unclear. Research has shown that livers of female rats develop a greater degree of intracellular acidosis during ischemia than males. This work explores the role of sex hormones in mediating this sex difference. METHODS: Subgroups of neutered female rats were given 17 beta-estradiol (E), progesterone (P), or combination (E+P). To compare the effects of female sex hormones in males, subgroups of intact and castrated males received 17 beta-estradiol. In vivo and ischemic liver biopsies were taken and analyzed for lactate and H. RESULTS: Although there was no effect of hormone therapy on baseline metabolic parameters, during ischemia compared to neutered females, livers from E females significantly (P<0.01) increased lactate by 56% and H+ by 71%, while E+P significantly increased only lactate (39%; P<0.05). Livers from neutered males given 17 beta-estradiol showed significantly greater (P<0.001) accumulation of lactate (80%) and H+ (79%). This was even shown in intact males, where despite a blunted response, 17 beta-estradiol, significantly (P<0.05) increased lactate by 39% and H+ by 25%. CONCLUSION: This study illustrates the mechanisms for the sex difference in the liver's metabolic response to ischemia are estrogen mediated, which is seen even in the presence of male hormones, thus offering one explanation for the lower liver transplant success when the donor is female.

Newborn hearts are at greater 'metabolic risk' during global ischemia--advantages of continuous coronary washout.

BACKGROUND: Altered metabolic responses of the newborn heart to ischemia, which may increase irreversible injury, may at least partially explain the greater morbidity and mortality experienced by some children undergoing congenital cardiac repair. The present study compared newborn heart metabolic responses to global ischemia with those of adult, and evaluated whether continuous coronary artery washout in the newborn heart during 'ischemia' could favourably affect these responses. METHODS: Adult (n=12) and newborn (n=12) pigs were anesthetized, and right ventricular biopsies were taken before global ischemia and at set intervals during ischemia. Another 12 newborns were subdivided into groups of nonperfused hearts and hearts receiving continuous perfusion. Time to onset of and time to peak of ischemic contracture were recorded. Biopsies were assayed for lactate, myocardial glycogen, glucose-6-phosphate and ATP. RESULTS: Newborn hearts were more sensitive to global ischemia than adult hearts, based on shorter time to onset of and time to peak of ischemic contracture, and had a significantly greater rate of ATP decline (P<0.01). This was due in part to a more rapid accumulation of lactate (P<0.05) and only a 50% use of glycogen, compared with 93% by adult hearts. Continuous washout of newborn hearts prevented lactate accumulation, allowing a 90% use of glycogen and delaying time to ischemic contracture by twofold. This was accompanied by lower levels of glucose-6-phosphate accumulation (P<0.05) and a threefold reduction in the rate of ATP decline. CONCLUSIONS: Significant differences in myocardial metabolism during ischemia in newborns compared with adults could predispose them to earlier ischemic injury, which can be eliminated by the removal of end products. Perfusion strategies taking these differences into account may further optimize pediatric myocardial protection and improve outcomes in newborn children undergoing cardiac procedures.

Are there ventricle-specific postnatal maturational differences in myocardial beta-adrenoceptors?

Newborn hearts have restricted functional reserve and variable responsiveness to inotropes that could be partly due to differences in myocardial beta-adrenoceptors (beta-AR). To clarify this issue, this study documented ventricle-specific changes in myocardial beta-AR density and affinity during postnatal maturation. In vivo left and right ventricle (LV and RV, respectively) biopsies were obtained from newborn (3-day-old, n = 11), immature (14-day-old, n = 7), and adult (n = 6) pigs. Total beta-AR density (B(max), fmol/g) and dissociation constant (K(d), pmol/L) were determined by radioligand binding with I125 iodocyanopindolol. Overall, beta-AR B(max) in the LV significantly decreased with maturation. Interestingly, newborn animal hearts (LV and RV) subdivided into 2 groups: an adult-like low K(d) group with low B(max) and a fetal-like high K(d) group with high B(max), which were significantly different from one another. The high K(d) newborn group also had significantly higher K(d) and B(max) than both immature and adult hearts. Newborns had similar Bmax but higher Kd in the LV than the RV, whereas immature and adult hearts did not have ventricular differences. During maturation, beta-AR density decreased, whereas LV beta-AR binding affinity increased. Variable beta-AR maturity was also identified immediately post partum, which could potentially explain the newborn heart's variable responsiveness to inotropes. The subset of newborn hearts with lower binding affinity (reduced responsiveness) could also contribute to the newborn heart's overall reduction in functional reserve.

Does hyperoxia affect glucose regulation and transport in the newborn?

OBJECTIVE: Hyperglycemia has been found to occur in children placed on cardiopulmonary bypass. Our laboratory demonstrated that hyperoxia plays a role in this hyperglycemic response and also occurs in the absence of cardiopulmonary bypass. The purpose of this study was to elucidate potential mechanisms underlying the hyperoxic-induced hyperglycemia by examining glucagon, insulin, and epinephrine, which are important in glucose regulation and skeletal and cardiac glucose transporters (GLUT1 and GLUT4), which facilitate glucose entry. METHODS: Three-day-old piglets were anesthetized, intubated, and ventilated to normoxia. Animals were then randomly allocated to either 5 hours of normoxia (n = 4) or hyperoxia (n = 6). Measurements of oxygen, blood glucose, plasma glucagon, insulin, and epinephrine levels were made. Total GLUT1 and GLUT4 content in cardiac and skeletal muscle was measured using Western blotting analysis. RESULTS: A sustained hyperglycemic response (P <.001) was seen throughout the 5-hour ventilatory period. A significant twofold elevation in glucagon levels (P <.001) and a threefold elevation (P <.003) in plasma insulin levels occurred, despite no significant changes in plasma epinephrine. Total GLUT1 and GLUT4 content were significantly reduced in skeletal muscle by 66% and 59%, respectively, while no significant changes occurred in cardiac muscle. CONCLUSION: This study demonstrates that significant elevations in glucagon and insulin and reductions in total skeletal muscle GLUT1 and GLUT4 content all contribute to hyperoxia-induced hyperglycemia seen in newborns. To optimize postoperative recovery of newborns, consideration should be given to the levels of oxygen used to avoid the potential development of insulin resistance and subsequent decrease in glucose entry.

Effect of hypertension on reproductive organ weights in various strains of rats.

The testes of Wistar Kyoto (WKY) and Spontaneously Hypertensive (SHR) rats have been shown to have differences in regional vascular resistance. In addition, myocardial hypertrophy tends to be more pronounced and occur more frequently in WKY and SHR female rats than in their male counterparts. Therefore, we sought to determine whether hypertension had any effect on reproductive organs and whether this effect was the same among strains. We removed the testes or uterus from Sprague Dawley (SD; n = 30 male and 30 female), WKY (n = 23 male and 16 female), and SHR (n = 27 male and 34 female) rats, weighed these animals and organs, and calculated the reproductive organ:body weight ratio (mean +/- 1 standard deviation x 10-3 g). The testes:body weight ratio of WKY (6.5 +/- 0.8) animals was significantly (P= 0.05) less than those of SD (7.6 +/- 1.1) and SHR (8.0 +/- 0.5) rats. The uterus:body weight ratio of the SD rats (1.7 +/- 0.4) was significantly (P = 0.05) less than those of WKY (2.4 +/- 0.6) and SHR (2.3 +/- 0.6) rats. Compared with WKY rats, male SD rats had a higher testes:body weight ratio whereas female SD rats had a lower uterus: body weight ratio. Whereas the SHR testes:body weight ratio was significantly higher than that of WKY rats, this effect was not seen for the uterus: body weight ratio. The effect of hypertension on reproductive organs should be taken into consideration when choosing a species, gender, or organ for study. As well, data compared across genders or strains must be evaluated carefully to ensure valid comparisons.

Preischemic administration of ribose to delay the onset of irreversible ischemic injury and improve function: studies in normal and hypertrophied hearts.

Compared with normal hearts, those with pathology (hypertrophy) are less tolerant of metabolic stresses such as ischemia. Pharmacologic intervention administered prior to such stress could provide significant protection. This study determined, firstly, whether the pentose sugar ribose, previously shown to improve postischemic recovery of energy stores and function, protects against ischemia when administered as a pretreatment. Secondly, the efficacy of this same pretreatment protocol was determined in hearts with pathology (hypertrophy). For study 1, Sprague-Dawley rats received equal volumes of either vehicle (bolus i.v. saline) or ribose (100 mg/kg) before global myocardial ischemia. In study 2, spontaneously hypertensive rats (SHR; blood pressure approximately 200/130) with myocardial hypertrophy underwent the same treatment protocol and assessments. In vivo left ventricular function was measured and myocardial metabolites and tolerance to ischemia were assessed. In normal hearts, ribose pretreatment significantly elevated the heart's energy stores (glycogen), and delayed the onset of irreversible ischemic injury by 25%. However, in vivo ventricular relaxation was reduced by 41% in the ribose group. In SHR, ribose pretreatment did not produce significant elevations in the heart's energy or improvements in tolerance to global ischemia, but significantly improved ventricular function (maximal rate of pressure rise (+dP/dt(max)), 25%; normalized contractility ((+dP/dt)/P), 13%) despite no change in hemodynamics. Thus, administration of ribose in advance of global myocardial ischemia does provide metabolic benefit in normal hearts. However, in hypertrophied hearts, ribose did not affect ischemic tolerance but improved ventricular function.

Body weight and food intake profiles are modulated by sex hormones and tamoxifen in chronically hypertensive rats.

Sex hormones and the selective estrogen receptor modulator tamoxifen affect food consumption and body weight in normotensive rats. This study investigated the effects of hormone manipulation and tamoxifen on weight gain and food intake in the presence of chronic systemic hypertension. Male and female spontaneously hypertensive rats (SHR) were either neutered or sham operated before puberty, and subgroups of neutered females received either estrogen replacement therapy (ERT) or tamoxifen at the age of 12 wk. Weekly body weight and food consumption were assessed, and food consumption was normalized to metabolic weight (g body(2/3)). Neutering reduced weight gain in males (P = 0.0001), but increased it in females (P < 0.0001). Both ERT and tamoxifen treatment prevented this increase in weight, with body weight dropping to levels of sham-operated rats for ERT, whereas rats given tamoxifen maintained greater body weights than sham-operated rats (P < 0.0001). This contrasts with previous work in normotensive females in which sham-operated and tamoxifen-treated females did not differ. Neutering reduced normalized food consumption relative to sham-operated rats in both males and females (P < 0.05). Although ERT returned it to normalized intakes of sham-operated rats, tamoxifen reduced normalized food consumption relative to that of both sham-operated and ERT groups. In hypertensive rats, body weight is modulated by sex hormones in both males and females, but in opposite directions. Both estrogen and tamoxifen exert immediate effects in females. Interestingly, the effect of tamoxifen on body weight appears to be greater in hypertensive than in normotensive rats.

Multiple in vivo liver biopsies using a freeze-clamping technique.

A multiple in vivo liver biopsy technique was developed to measure labile metabolites (creative phosphate [CP], ATP, lactate) without interfering with normal perfusion and metabolism. Anesthetized rats (n = 7) had a midline abdominal incision done to expose the liver. Four biopsies were taken across 20 min. Bleeding was controlled by a small, nontraumatic clamp proximal to the biopsy. Prefrozen dressing forceps grasped the liver and scissors cut the biopsy, which was frozen in liquid nitrogen. Bleeding was minimal and hemodynamic stability was preserved. This technique has few complications, bleeding is easily controlled, and it allows for large multiple biopsies, which give reliable metabolic data and can be consistently taught.

Special Feeding and Care of Senescent Spontaneously Hypertensive Rats.

This study investigated the impact of feeding methods on body weight of senescent female spontaneously hypertensive rats (SHRs) and showed that supplementing powdered feed was useful as they approached heart failure at 22 to 23 months of age. SHRs are genetically predisposed to systemic hypertension and will, with age, progress into complete heart failure resulting in death. Close to the time of heart failure, some rats experienced a loss of appetite and weight loss. It was postulated that either elevated blood pressure, age-associated health issues, or the effort required to access pelleted food prevented the rats from eating properly, resulting in malnutrition and weight loss. As they aged, the rats benefited from the addition of powdered food to ensure that body weights remained stable and to prevent malnutrition that could lead to premature death. Animals were fed commercially available rat chow pellets until they showed persistent signs of weight loss or a lack of interest in their food. At that time, the rats were also given powdered rat chow in shallow bowls to facilitate the eating and the digestion of their food. The rats were weighed weekly to confirm they consumed sufficient calories daily and to ensure that the change to the powdered chow was having the desired effect. Prior to being fed the powdered rat chow, the rats had shown signs of progressive weight loss. After starting the powdered chow, the rats either maintained or gained weight. This study shows that as the female SHR matures, special care and handling is key to maintaining body weights and good health. With only modest changes in routine (i.e., powdered food) and an attentive eye on the rats' daily activities, it was possible to maintain these senescent female SHRs in a healthy condition until the termination of the study or onset of heart failure.