αCaMKII controls the establishment of cocaine's reinforcing effects in mice and humans.

Although addiction develops in a considerable number of regular cocaine users, molecular risk factors for cocaine dependence are still unknown. It was proposed that establishing drug use and memory formation might share molecular and anatomical pathways. Alpha-Ca(2+)/calmodulin-dependent protein kinase-II (αCaMKII) is a key mediator of learning and memory also involved in drug-related plasticity. The autophosphorylation of αCaMKII was shown to accelerate learning. Thus, we investigated the role of αCaMKII autophosphorylation in the time course of establishing cocaine use-related behavior in mice. We found that αCaMKII autophosphorylation-deficient αCaMKII(T286A) mice show delayed establishment of conditioned place preference, but no changes in acute behavioral activation, sensitization or conditioned hyperlocomotion to cocaine (20 mg kg(-1), intraperitoneal). In vivo microdialysis revealed that αCaMKII(T286A) mice have blunted dopamine (DA) and blocked serotonin (5-HT) responses in the nucleus accumbens (NAcc) and prefrontal cortex after acute cocaine administration (20 mg kg(-1), intraperitoneal), whereas noradrenaline responses were preserved. Under cocaine, the attenuated DA and 5-HT activation in αCaMKII(T286A) mice was followed by impaired c-Fos activation in the NAcc. To translate the rodent findings to human conditions, several CAMK2A gene polymorphisms were tested regarding their risk for a fast establishment of cocaine dependence in two independent samples of regular cocaine users from Brazil (n=688) and Switzerland (n=141). A meta-analysis across both samples confirmed that CAMK2A rs3776823 TT-allele carriers display a faster transition to severe cocaine use than C-allele carriers. Together, these data suggest that αCaMKII controls the speed for the establishment of cocaine's reinforcing effects.

The reliability of activated partial thromboplastin time methods and the relationship to lipid composition and ultrastructure.

Wide variations in procoagulant properties, lipid composition and ultrastructure of five commonly used APTT methods have been demonstrated. Performance of the methods with a range of coagulation abnormalities has been ranked. Most of the reagents obtained a high score with one or more defects, but a low score with others. A consistent good ranking throughout was only observed with one reagent. The number of significant correlations between the reagents' procoagulant activities and lipid content confirms the view that the performance of an APTT method is largely dependent upon its lipid composition. Marked differences in concentration and distribution of phospholipids, fatty acids and neutral lipids were evident. The importance of the concentration of phosphatidyl serine in regulating the procoagulant activity of an APTT method has been demonstrated. Electron microscopy provides evidence of the contrasting composition of the reagents from the more discrete uniform liposomes present in the more reliable reagents, to more ill-defined components present in those reagents which performed less well. The study highlights the need for standardisation of the APTT.

Lipid class composition and heparin sensitivity in the activated partial thromboplastin time.

In an APTT reagent, prepared from purified lipids, the role of phosphatidyl serine (PS) in determining the sensitivity of the APTT test system to measurement of the effect of heparin in plasma has been evaluated. As the concentration of PS decreases sensitivity to heparin increases but procoagulant activity decreases. Dilution of the test liposome over a wide range (1 g/l to 30 mg/l) had a minimal effect on the clotting time. At levels below 30 mg/l, however, the amount of total lipid appeared to be rate limiting; a loss of procoagulant activity being paralleled by an increase in heparin sensitivity. Phosphatidyl inositol (PI) was not a satisfactory substitute for PS in the APTT method studied. The degree of unsaturation of test liposomes appeared to have no effect on either procoagulant activity or sensitivity to heparin at the lipid concentration employed. In the light of these findings, a more critical appraisal of the phospholipid components of APTT reagents should facilitate the development of more reliable reagents for heparin control. A further benefit of this type of approach should be a reduction in the acknowledged wide variations in sensitivity to heparin which exist between available APTT reagents.

The use of Vacutainer tubes for collection and storage of blood for coagulation testing.

A study has been performed to evaluate the suitability of Vacutainer tubes in blood specimen collection for coagulation tests and to compare them with the conventional syringe technique employed in UK hospitals. Blood was collected from healthy volunteers, an ante-natal group and patients on long-term oral anticoagulants. Samples were stored at two different temperatures; 4 degrees C and ambient room temperature (RT). Prothrombin times, factor VII assays and APTT were performed at baseline and after 2 h and 4 h storage. There was significant activation of the extrinsic system in the blood samples collected by Vacutainer when stored at 4 degrees C which became more significant on prolonged storage. The effect was less pronounced when the Vacutainer tubes were stored at RT. In contrast, the blood collected by the syringe method did not show these changes with the exception of the ante-natal specimens where a lesser degree of activation than in the Vacutainer tubes was observed after 4 h at 4 degrees C. The activation of the Vacutainer samples at 4 degrees C is considered undesirable and could be of clinical significance in oral anticoagulant dosage.

An assessment of an amidolytic assay for factor VII in the laboratory control of oral anticoagulants.

A comparison has been made between the prothrombin time test using British Comparative Thromboplastin (BCT) and a chromogenic substrate assay for factor VII in the assessment of laboratory control of oral anticoagulants in short-term and long-term patients. Opportunity was also taken to compare the findings with parallel results obtained with the venous Thrombotest technique and a specific clotting assay for factor VII. There was good agreement between the amidolytic factor VII assay, using a method modified from Seligsohn et al (1978) with the Quick test using BCT and Thrombotest in 60 long-term patients. Tests in 53 patients within the first 3 weeks of starting oral anticoagulant administration gave less satisfactory agreement between the above amidolytic method and the conventional tests. In contrast, there was a good correlation between the two conventional tests in both groups and also between the clotting and amidolytic factor VII method. Although the results are an improvement on previous, less satisfactory correlations between the BCT prothrombin time method and amidolytic assays for factor II and X, the present study indicates the limitations of a specific clotting assay versus a broad spectrum extrinsic clotting test in oral anticoagulant control. While not warranting the routine use of the chromogenic assay for factor VII in place of the prothrombin time using BCT, the factor VII amidolytic assay offers a limited but dependable guide to dosage in long-term patients. The complexity of the technique in its present form militates against its adoption for routine anticoagulant control in hospital laboratories.