[Identification of a Novel Calcium (Ca

Calcium (Ca^(2+))-activated chloride channel accessories (CLCAs) are putative anion channel-related proteins with diverse physiological functions. Exploring CLCA diversity is important for prediction of gene structure and function. In an effort to identify novel CLCA genes in Xenopus laevis, we successfully cloned and characterized a Xenopus laevis cDNA predicted to encode the xCLCA3 gene. Cloning of xCLCA3 was achieved by computational analysis, rapid amplification of cDNA ends (RACE), and a tissue distribution analysis by semi-quantitative reverse transcription (RT) PCR or real-time PCR. We obtained a 2958 bp xCLCA3 cDNA sequence with an open reading frame encoding 943 amino acids. According to the primary structure analysis, xCLCA3 contains a predicted signal sequence, multiple sites of N-linked (N-) glycosylation, N-myristoylation, PKA, PKC, and casein kinase II phosphorylation sites, five putative hydrophobic segments, and the HExxH metalloprotease motif. Additionally, the transmembrane prediction server yielded a preserved N-terminal CLCA domain and a von Willebrand factor type A domain with one transmembrane domain in the C-terminal region. Expression analysis showed that xCLCA3 is expressed in a number of tissues, with strong expression in the brain, colon, small intestine, lung, kidney, and spleen, and poor expression in the heart and liver. These results suggest that xCLCA3 may be a candidate CLCA family member as well as a metalloprotease, rather than just an ion channel accessory protein.

Fish oil, essential fatty acids, and hypertension.

A proper balance between the n-3 and n-6 series of essential fatty acids (EFAs) is essential for homeostasis and normal growth in humans. Dietary supplement with fish oil and related n-3 EFAs has been used to study their antihypertensive property in animals and humans with borderline and essential hypertension. In the animal models, chronic treatment of young animals generally only attenuated the development of hypertension. In animals with hypercholesterolemia, n-3 EFA supplement increased the incidence of atherosclerosis. In humans, chronic treatment with fish oil only produced a small reduction in blood pressure. The concerns are that the high dose of fish oil may interfere with the control of blood glucose in diabetic patients, and may cause prolonged bleeding in surgical patients. Studies on the animal models of hypertension showed that n-6 EFAs are more effective than n-3 EFAs in lowering and normalizing the blood pressure of these animals, probably through the production of tissue prostaglandins, which favour vasodilation. The antihypertensive effect of the n-6 EFAs in humans is not well known, because there are only a few studies, usually involving a very small number of patients. A possible side effects of n-6 EFAs for concern is that they might stimulate tumour development. A careful examination of these risk factors is needed before any recommendation can be made concerning the use of EFAs for the control of hypertension for humans.

Measurement of both cyclic [3H]AMP and cyclic [3H]GMP in cultured vascular smooth muscle cells labeled with [3H]hypoxanthine: use in studies of cardiovascular drugs.

We describe a method for prelabeling cultured vascular smooth muscle cells that permits rapid and accurate measurements of changes in the amounts of cyclic AMP and of cyclic GMP in 5 x 10(5) cells. This procedure utilizes [3H]hypoxanthine to radiolabel both the adenine and guanine nucleotide pools and simple column chromatographic steps to isolate and separate the 3H-labeled cyclic nucleotides. The application of the method to studies of the actions of cardiovascular drugs on vascular smooth muscle cells is illustrated by measurements of the effects of isoproterenol, nitroprusside, and inhibitors of cyclic AMP phosphodiesterases on the cyclic nucleotide levels in these cells. If required, the mass amounts of cyclic AMP and cyclic GMP present could be determined by measurement of the specific radioactivities of the precursor [3H]ATP and [3H]GTP, respectively. The cyclic nucleotide values calculated by the latter method were almost identical to those obtained with larger numbers of cells using commercially available radioimmunoassays, thus validating the prelabeling assays. The method described should be applicable to any type of cultured cell that can utilize [3H]hypoxanthine to replenish its ATP and GTP pools.

Discrimination of systolic blood pressure.

The ability of humans to discriminate systolic blood pressure (BP) was investigated in two experiments. In Experiment 1, 14 normal subjects were asked to make estimates of their systolic BP while performing both BP-elevating and BP-lowering tasks. They were given intermittent feedback throughout all 10 45-min sessions. Results indicated significant correlations and small absolute differences between estimated and measured BP for all subjects in almost all sessions. Experiment 2, undertaken 6 months after Experiment 1, assessed whether estimation accuracy by subjects who had available both external and interoceptive cues surpassed that of subjects which access to external cues only. Three subjects from the original group who showed consistently high motivation, and who improved in accuracy across the 10 sessions in the previous experiment, made estimates of BP while performing novel tasks with no feedback. Correlations between estimated and measured BP remained high for 2 of the 3. These results were compared with the accuracy of control subjects (3 for each experimental subject) who were asked to estimate experimental subjects' BP using only the cognitive information available to the experimental subjects. Control subjects also had high correlations between their estimates and the experimental subjects' measured BP but at lower levels than two experimental subjects. These findings are discussed in relation to subjects' possible use of interoceptive information.

Blood pressure tracking systems and their application to biofeedback.

Two automatic electronic systems for tracking diastolic or systolic blood pressure (BP) and their use in biofeedback experiments are described. Both systems are based on the use of an arm cuff and the recording of Korotkoff sounds with a microphone. An electronic control system receives input from the microphone and provides output to several solenoid valves that control compressed airflow to the cuff. For one of the tracking systems we devised, the cuff is alternately inflated for 45 sec and deflated for 45 sec. During inflation, small increments and decrements in cuff pressure, based on the occurrence of Korotkoff sounds, are used to track BP. The other tracking system is slower but provides more continuous measurements: The cuff is alternately inflated to approximately the BP level for 5 sec and then deflated for 5 sec. During each inflation, the number of Korotkoff sounds is recorded by the system and according to this number, the amount of inflation on the next trial is automatically adjusted so that the cuff pressure follows BP. Both systems have been used successfully in biofeedback applications.