Measurement of methemoglobin in neonatal samples containing fetal hemoglobin.

Because of the potential for methemoglobinemia during nitric oxide therapy in newborns, methods are needed to accurately quantify methemoglobin (MetHb) in the presence of the high concentrations of fetal hemoglobin (Hb F), bilirubin, and lipids seen in these patients. Spectral differences between fetal and adult Hbs invalidate assumptions of conventional multiwavelength Hb photometry, so we evaluated an "overdetermined" system (Ciba-Corning Model 270), in which absorbances at seven wavelengths are measured to quantify four Hb derivatives. Adult and umbilical cord blood (Hb F 96%) samples were prepared to contain known MetHb fractions. Measured MetHb was linear in cord blood to > or = 15% MetHb. Within-run precision (CV) was < 2.2% (n = 10) at each of seven MetHb fractions between 5% and 100%. Measured (y) and expected (x) MetHb fractions in cord blood were in good agreement (y = 1.0200x + 0.100, Sylx = 0). Added bilibrubin (200 mg/L serum) and lipid (30 g/L) did not interfere. No significant differences were seen for adult and cord blood samples with identical MetHb fractions (P = 0.72), whereas a significant difference was noted with an exactly determined system (P = 0.0033). At clinically relevant MetHb fractions (< 15%), a trend towards increased values in cord blood was noted with an exactly determined system (y = 1.0520x + 0.7600). We conclude that this overdetermined system measures MetHb accurately in samples from patients with large concentrations of Hb F.

Levels of angiotensin and molecular biology of the tissue renin angiotensin systems.

The cloning of renin, angiotensinogen and angiotensin converting enzyme genes have established a widespread presence of these components of the renin-angiotensin system in multiple tissues. New sites of gene expression and peptide products in different tissues has provided strong evidence for the production of angiotensin independently of the endocrine blood borne system. In addition, the cloning of the angiotensin receptor (AT1) gene has confirmed the widespread distribution of angiotensin and suggested new functions for the peptide. This review of various tissues shows the variation in gene expression between tissues and angiotensin levels, and the fragmentary state of our knowledge in this area. As yet we cannot state that the gene expression of the substrates, enzymes and peptide products are involved in a single cell synthesis. This is not so much evidence against a paracrine function for tissue angiotensin, as lack of detailed, accurate intracellular information. The low abundance of renin in brain, spleen, lung and thymus compared to kidney, adrenal, heart, testes, and submandibular gland may suggest that there are both tissue renin-angiotensin systems (RAS) and nonrenin-angiotensin systems (NRAS). The NRAS could function through cleavage of angiotensinogen by serine proteinases such as tonin and cathepsin G to form Ang II directly. Although much angiotensinogen is extracellular and could therefore be a site of synthesis outside of the cell, intracellular angiotensinogen in a NRAS process could produce Ang II intracellularly without requiring extracellular conversion of Ang I to Ang II by ACE. In summary, renin mRNA is found in high concentrations in kidney, adrenal and testes and decreasing lower concentrations in ovary, liver, brain, spleen, lung and thymus. Angiotensinogen mRNA is found in the following tissues in descending order of abundance: liver, fat cells, brain (glial cells), kidney, ovary, adrenal gland, heart, lung, large intestine and stomach. It is debatable whether angiotensinogen and renin mRNA are expressed in blood vessels. The evidence that is lacking for a paracrine function of angiotensin is a complete description of the intracellular molecular synthesis and release of Ang II from single cells of promising tissues. Such tissues, SMG, ovary, testes, adrenal, pituitary and brain (neurons and glia) are potent sources of RAS components for future studies. Although the evidence for a paracrine function of angiotensin II is incomplete, it is an important concept for progressing toward the understanding of tissue peptide physiology and the significance of their gene regulation.

The role of angiotensin, AT1 and AT2 receptors in the pressor, drinking and vasopressin responses to central angiotensin.

Angiotensin II (Ang II) given centrally produces an increase in blood pressure and motivation to drink. The physiological mechanisms that mediate the pressor response include release of vasopressin (AVP) and activation of the sympathetic nervous system. Using 2 new Ang II receptor antagonists, we were able to investigate the role of AT1 or AT2 receptors in mediating these effects. Adult male Sprague-Dawley rats were cannulated in the lateral ventricle and 5 days later catheterized in the carotid artery for blood pressure measurements. All experiments were carried out in conscious rats. Three treatments were given intraventricularly (i.v.t.), in 2 microliters artificial cerebrospinal fluid (ACSF) at 30 min intervals: (1) 50 ng Ang II, (2) 0.7 micrograms AT1 antagonist Losartan or 7.0 micrograms AT2 antagonist PD123177, followed by 50 ng Ang II, and (3) 50 ng Ang II, to test for recovery. Blood pressure and drinking measurements were recorded. Also, blood samples for assay of AVP were drawn at 1 or 3 min post-injection in 2 separate groups of rats. We found that both Losartan and PD123177 significantly reduced release of AVP to Ang II 1 min post-injection. Losartan significantly blocked the pressor response (P less than 0.001), while PD123177 had no significant effect. Drinking was also antagonized by Losartan (P less than 0.05) and reduced (n.s.) by PD123177. The results suggest that the pressor response to Ang II (i.v.t.) is predominantly AT1 mediated, while the drinking and AVP responses may be mediated by both receptor subtypes.

Blood volume changes of the anaesthetized rat to a large burn or mock burn injury measured using a continuous measuring circuit.

A scald burn injury of 40-60 per cent body surface area was applied to the sodium pentobarbitone anaesthetized rat. The cardiac output of the burn injury rats fell from 39.5 +/- 2.1 to 27.7 +/- 1.5 ml/min (P less than 0.001) while heart rate, mean arterial blood pressure and central venous pressures were little changed but the total peripheral resistance rose significantly. Cardiovascular function was unchanged in control, mock-burned rats. The blood volume was measured continuously using an extracorporeal circuit and was observed to fall significantly by 0.78 +/- 0.36 ml in 5 min (P less than 0.05) and 1.65 +/- 0.38 ml in 60 min (P less than 0.001) after burn injury. The falls in blood volume and cardiac output were virtually simultaneous, and had occurred by 5 min. It is proposed however that this fall in blood volume is not sufficient to cause the observed changes in cardiac output and that additional factors such as cardiac impairment may be responsible for these changes postburn.

Effect of allopurinol or superoxide dismutase plus catalase on cardiovascular function after burn injury in the anaesthetized rat.

A scald burn injury was applied to 22-23 per cent of the body surface area of anaesthetized rats. The cardiac output was 38-60 per cent lower in the burn injury group than in the control group; heart rate and mean arterial blood pressure were only slightly affected, but burn injury caused a significant increase in total peripheral resistance. The involvement of oxygen-free radicals in this immediate fall in cardiac output was investigated. Pretreatment with a blocker of free radical production, allopurinol or the infusion of the free radical scavengers superoxide dismutase plus catalase caused no cardiovascular improvement, suggesting that oxygen free radicals are not involved in the fall in cardiac output after burn injury. Allopurinol treatment, however, prevented the rise in total peripheral resistance seen after burn injury.