Donald H. Namm

Species specificity of the platelet responses to 1-0-alkyl-2-acetyl-sn-glycero-3-phosphocholine.

A semi-synthetic 1-0-alkyl-2-acetyl-sn-glycero-3-phosphocholine (alkylacetyl-GPC) was shown to be highly species selective in its capacity to cause platelet aggregation and serotonin release. No effects were elicited on the rat or mouse platelets while platelets from human, dog, cat, rabbit, guinea pig and horse were highly sensitive to alkylacetyl-GPC. The hypotensive activity in the rat was not associated with thrombocytopenia. Preliminary evidence suggested that the inability of platelets of the rat and mouse to respond to alkylacetyl-GPC was not due to a difference in plasma inactivation of the substance but due to a difference in platelet responsiveness per se. The data also support the concept that the potent hypotensive property of this substance readily observed in the rat, is a result of an effect which is platelet-independent.

Synthesis of prostaglandin 6-keto F1α by cultured aortic smooth muscle cells and stimulation of its formation in a coupled system with platelet lysates

Lysed aortic smooth muscle cells, when incubated with [14C] arachidonate, synthesized only one radioactive product, which was identified as 6-keto-PGF1alpha. Formation of this product from smooth muscle cell lysates was stimulated when human platelet extracts were added to the system, and further stimulation was observed when imidazole, a selective inhibitor of thromboxane synthesis, was added to this coupled system. These observations indicate that the cyclooxygenase of the smooth muscle cells was rate-limiting, that the prostacyclin synthetase of these cells can utilize endoperoxides produced by platelets, and that blocking of thromboxane synthesis might, under certain conditions, shunt arachidonate metabolism toward prostacyclin formation.

Characterization of the thrombin-induced contraction of vascular smooth muscle

Purified human alpha-thrombin induced a sustained contraction of isolated rabbit aorta and dog coronary arteries. These vascular tissues also exhibited a refractoriness towards a second thrombin exposure. The extent of tachyphylaxis exhibited by the aorta correlated with the initial concentration of thrombin and the length of time the tissue was exposed to thrombin. The thrombin-induced contraction in the aorta was not blocked by phospholipase or cyclooxygenase inhibitors, but it was inhibited in the presence of hirudin, heparin, nitroglycerin, and nitroprusside. Nitroglycerin, nitroprusside, and hirudin also inhibited the contraction in the dog coronary artery. Ca++ channel blockers did not inhibit the thrombin-induced contraction in the coronary artery, although a small inhibition was observed in Ca++-free media. In both tissues, equivalent contractile responses were obtained using equimolar quantities of beta-, tetranitromethane-, and alpha-thrombin, even though the latters coagulant activity was 30-40 times that of the modified thrombins. However, if the catalytic activity of thrombin was inhibited by modification with Tos-Lys-CH2Cl, hirudin, or heparin/antithrombin III, the vasoconstrictor activity was also lost. These studies suggest that alterations of the fibrinogen-binding site do not affect the contractile activity of thrombin. The contraction may be the result of a proteolytic interaction of the active site of the enzyme with vascular smooth muscle.

Occurrence and function of cyclic nucleotides in blood vessels.

The literature concerned with studies of the occurrence and function of the cyclic nucleotides in blood vessels is reviewed. Emphasis is placed on the critical evaluation of the evidence which relates to the hypothesis that cyclic nucleotides meditate the effects of drugs and neurotransmitters on vascular contractility. The hypothesis that cyclic AMP mediates vasodilation, especially that induced by beta-adrenergic relaxation, is supported by many experimental approaches, but it is concluded that the evidence remains unconvincing based on the criteria established for such a mediator role. Possible sites of action of cyclic AMP are discussed. The demonstrated action of cyclic AMP on vascular membrane electrophysiology and calcium ion pumps are reviewed as possible causes of relaxation. The role of both nucleotides in vascular disease, especially hypertension is discussed. Finally the needs for further research in this area are suggested.

A sensitive analytical method for the detection and quantitation of adenosine in biological samples.

Abstract A new method for the measurement of adenosine in biological materials has been developed. The method is based on the combined principles of isotope dilution and enzymatic catalysis using a highly specific adenosine kinase isolated from rat heart. By differential centrifugation and gel filtration, this adenosine kinase was obtained free of adenosine deaminase and other enzymes which would have been a source of error in the use of this enzyme in the adenosine assay. The cardiac adenosine kinase was shown to be highly specific and to exhibit an apparent K m for adenosine of 0.35 μ M . Using this enzyme, unknown quantities of adenosine could be detected by measuring the effect of their addition on the conversion of radioisotopic adenosine to 5′-AMP in the kinase reaction. In this procedure, as little as 20 pmoles of adenosine could be detected. To test the applicability of the assay, measurements of the tissue content of this nucleoside were made in samples of dog and rat hearts frozen in situ under control, hypoxic, or ischemic conditions. The assay has several advantageous features when compared to other existing methods used to measure adenosine: a minimum of sample preparation is required before the actual assay procedure; many samples can be processed per day by a single operator; single determinations can be done on as little as 5 μl of sample, and the specificity of the assay can be readily checked by treatment of samples with adenosine deaminase.

Generation of a Vasoactive Substance in Human Plasma during Coagulation

A stimulant of vascular smooth muscle contraction was generated in fresh, citrated human plasma during activation of the clotting system. Plasma, exposed briefly to thromboplastin and Ca++, induced a contraction of isolated rabbit aorta and dog coronary arteries that was slow in development and persisted after washout. The contractile activity was not blocked by phenoxybenzamine, atropine, or angiotensin inhibitor, but was blocked when heparin or hirudin was incubated with the plasma. The contractile stimulant produced in the plasma was short-lived (less than 3 min) and paralleled the appearance of thrombin in plasma. Purified human alpha-thrombin also induced a sustained contraction in these blood vessels that was not inhibited by phenoxybenzamine, atropine, or angiotensin inhibitor, but was blocked by hirudin. Partial relaxation of the thrombin-treated blood vessel was achieved by the addition of heparin. These results suggest that this vasoactive component of thromboplastin-activated human plasma is alpha-thrombin. Because of its potent and persistent effects, thrombin-induced vasospasm may be an important mechanism in the etiology of ischemic heart disease.

Incorporation of the Terminal Phosphate of ATP into Membranal Protein of Rabbit Cardiac Sarcoplasmic Reticulum: CORRELATION WITH ACTIVE CALCIUM TRANSPORT AND STUDY OF THE EFFECTS OF CYCLIC AMP

The transfer of the terminal phosphate of ATP to a material from the sarcoplasmic reticulum of cardiac muscle which can be precipitated by trichloroacetic acid was studied, and the relationship of this biochemical event to active Ca2+ transport was examined. A component of the phosphoryl transfer reaction was stimulated specifically by Ca2+. Both the phosphorylation of the reticulum and the active sequestering of Ca2+ were proportional to the Ca2+ concentration between 10−7 and 10−5M. The time course of both phenomena was similar. These and other observations relating Ca2+-dependent phosphorylation of the membrane of the sarcoplasmic reticulum with the activesequestering of Ca2+ suggest that the phosphoryl transfer reaction may represent the formation of a carrier system which facilitates the inward flux of Ca2+ against a concentration gradient. Cyclic AMP, theophylline, or the combination of these two agents did not influence the rate or extent of the phosphorylation of the sarcoplasmic reticulum. These results have led us to question the hypothesis that the cardiac sarcoplasmic reticulum is the site of action of cyclic AMP in its role as a mediator of cardiac inotropic effects.

Fitting the strategic planning process to organization: A clinical study of Burroughs Wellcome Co.

Abstract Strategic planning is art as well as science. Clinical analysis of Burroughs Wellcome Co. provides us with insight and understanding of the process. A number of themes are developed in this paper, with implementation as the first order of concern, since it is the driving mechanism for success. To begin at the end is mandatory if strategic planning is to be real for the organization ; questions are more important than answers. Rhetorically, good answers require good questions. This paper demonstrates that appropriate questions are necessary to keep the planning process on track. Initially, queries on strengths and weaknesses generated little response. However, questions such as “What do we do well?” and “What can we do better?” led to substantive issues. Later, the questions were refined to make stakeholders of the major organizational players. Some suggestions for strategic planning: the questions and the procedures must fit the organizational style; balance and cohesion must be considered and maintained at each step; and finally, strategic planning must be serious business, and the chief executive officers leadership is mandatory.