Relative contributions of connexins 40 and 43 to atrial impulse propagation in synthetic strands of neonatal and fetal murine cardiomyocytes

Atrial tissue expresses both connexin 40 (Cx40) and 43 (Cx43) proteins. To assess the relative roles of Cx40 and Cx43 in atrial electrical propagation, we synthesized cultured strands of atrial myocytes derived from mice with genetic deficiency in Cx40 or Cx43 expression and measured propagation velocity (PV) by high-resolution optical mapping of voltage-sensitive dye fluorescence. The amount of Cx40 and/or Cx43 in gap junctions was measured by immunohistochemistry and total or sarcolemmal Cx43 or Cx40 protein by immunoblotting. Progressive genetic reduction in Cx43 expression decreased PV from 34+/-6 cm/sec in Cx43(+/+) to 30+/-8 cm/sec in Cx43(+/-) and 19+/-11 cm/sec in Cx43(-/-) cultures. Concomitantly, the cell area occupied by Cx40 immunosignal in gap junctions decreased from 2.0+/-1.6% in Cx43(+/+) to 1.7+/-0.5% in Cx43(+/-) and 1.0+/-0.2% in Cx43(-/-) strands. In contrast, progressive genetic reduction in Cx40 expression increased PV from 30+/-2 cm/sec in Cx40(+/+) to 40+/-7 cm/sec in Cx40(+/-) and 45+/-10 cm/sec in Cx40(-/-) cultures. Concomitantly, the cell area occupied by Cx43 immunosignal in gap junctions increased from 1.2+/-0.9% in Cx40(+/+) to 2.8+/-1.4% in Cx40(+/-) and 3.1+/-0.6% in Cx40(-/-) cultures. In accordance with the immunostaining results, immunoblots of the Triton X-100-insoluble fraction revealed an increase of Cx43 in gap junctions in extracts from Cx40-ablated atria, whereas total cellular Cx43 remained unchanged. Our results suggest that the relative abundance of Cx43 and Cx40 is an important determinant of atrial impulse propagation in neonatal hearts, whereby dominance of Cx40 decreases and dominance of Cx43 increases local propagation velocity.

Effects of cerebral perfusion pressure and increased fraction of inspired oxygen on brain tissue oxygen, lactate and glucose in patients with severe head injury

OBJECTIVE: The purpose of the study was to measure the effects of increased inspired oxygen on patients suffering severe head injury and consequent influences on the correlations between CPP and brain tissue oxygen (PtiO2) and the effects on brain microdialysate glucose and lactate. METHODS: In a prospective, observational study 20 patients suffering severe head injury (GCS< or =8) were studied between January 2000 and December 2001. Each patient received an intraparenchymal ICP device and an oxygen sensor and, in 17 patients brain microdialysis was performed at the cortical-subcortical junction. A 6 h 100% oxygen challenge (F IO2 1.0) ( Period A) was performed as early as possible in the first 24 hours after injury and compared with a similar 6 hour period following the challenge ( Period B). Statistics were performed using the linear correlation analysis, one sample t-test, as well as the Lorentzian peak correlation analysis. RESULTS: F IO2 was positively correlated with PtiO2 (p < 0.0001) over the whole study period. PtiO2 was significantly higher (p < 0.001) during Period A compared to Period B. CPP was positively correlated with PtiO2 (p < 0.001) during the whole study. PtiO2 peaked at a CPP value of 78 mmHg performing a Lorentzian peak correlation analysis of all patients over the whole study. During Period A the brain microdialysate lactate was significantly lower (p = 0.015) compared with Period B. However the brain microdialysate glucose remained unchanged. CONCLUSION: PtiO2 is significantly positively correlated with F IO2, meaning that PtiO2 can be improved by the simple manipulation of increasing F IO2 and ABGAO2. PtiO2 is positively correlated with CPP, peaking at a CPP value of 78 mmHg. Brain microdialysate lactate can be lowered by increasing PtiO2 values, as observed during the oxygen challenge, whereas microdialysate glucose is unchanged during this procedure. Extension of the oxygen challenge time and measurement of the intermediate energy metabolite pyruvate may clarify the metabolic effects of the intervention. Prospective comparative studies, including analysis of outcome on a larger multicenter basis, are necessary to assess the long term clinical benefits of this procedure.

Inhalation anesthesia of rats: influence of the fraction of inspired oxygen on limb ischemia/reperfusion injury

Inhalation anesthesia with isoflurane is a well-established and safe method used in small laboratory animals. In most cases oxygen is used as a carrier gas for isoflurane, but room air or mixtures of oxygen with air or nitrous oxide are also being used. Anesthesia is therefore administered using different fractions of inspired oxygen (FiO2), and this may have consequences for the outcome of experiments. The aim of the present study was to investigate the influence of FiO2 on rat hind limb ischemia/reperfusion injury and to refine the used inhalation anesthesia. Male Wistar rats were subjected to 3.5 h of ischemia and 2 h of reperfusion, and divided into three groups according to FiO2 in the O2/air/isoflurane anesthesia gas mixture: 40%, 60%, and 100% O2. Normal, healthy rats were used as controls. Muscle edema and creatine kinase MM, a marker for myocyte necrosis, were significantly increased with 40% FiO2 as compared with 100% FiO2 (P<0.05). Partial pressure of oxygen, oxygen saturation, and oxyhemoglobin were significantly higher in the 100% O2 group as compared with 40% O2. No significant differences were detected for other parameters, such as the oxidative stress markers malondialdehyde and superoxide dismutase. We conclude that a refined inhalation anesthesia setting using 40% FiO2, reflecting more or less the clinical situation, leads to a more severe and more physiologically relevant reperfusion injury than higher FiO2. Oxidative stress did not correlate with FiO2 and seemed to have no influence on reperfusion injury.