Rectal bioavailability of lidocaine in man: partial avoidance of "first-pass" metabolism.

It is often speculated that after rectal administration drugs will enter the systemic circulation without first passing through the liver, because at least the lower hemorrhoidal veins are not connected to the portal system. To test this hypothesis, the systemic availability of the high-clearance drug lidocaine was investigated in 6 healthy subjects following administration of 200 mg intravenous, 300 mg oral, and 300 mg rectal lidocaine in a balanced crossover design. Plasma and whole blood concentrations of lidocaine were measured by capillary gas chromatography. The mean rectal systemic availability was higher than the oral: 63% vs 31% (whole blood) and 71% vs 34% (plasma). The elimination half-lifes (t1/2els) lidocaine were about the same intravenously and orally, whereas these were slightly longer after rectal administration. The oral and rectal investigations were repeated in the same panel of volunteers about 6 mo later. The mean rectal systemic availability, based on plasma concentrations, was then 67% vs 27% orally. Intraindividual variability was rather small, indicating that oral and rectal bioavailability of lidocaine is reproducible in individuals. An equation was derived for the calculation of the fraction of the dose given rectally that bypasses the liver after absorption which is slightly more than half the dose, assuming that dose is 100% absorbed. This investigation indicates that in principle it is possible to avoid, at least partly, drug loss caused by "first-pass" metabolism by giving the drug rectally.

The appearance of donor heparin in the recipient after reperfusion of a liver graft.

The release of heparin has been mentioned as one of the causes of hypocoagulability after reperfusion of the liver graft. It has been ascribed to endogenous heparin released from the donor liver or to exogenous heparin in the preservation fluid that is released into the recipient after sequestration into the graft during preservation. The aim of this study was to investigate whether systemic administration of heparin to the donor before the hepatectomy contributes to the appearance of heparin in the recipient after reperfusion. We studied 20 patients undergoing an auxiliary heterotopic liver transplantation; 15 donors had received heparin immediately before circulation arrest (median 300 IU/kg body weight), but 5 had not. The thrombin time (TT), activated partial thromboplastin time (aPTT), and heparin neutralization test were determined at several intervals during the transplantation.

Effects of sodium azide on platelet function.

Sodium azide in low concentrations (0.1-10 micrometer) was found to have inhibitory effects on human platelet function. Primary aggregation induced by ADP, epinephrine, thrombin and the ionophore A 23187 was decreased. To evaluate the effect of azide apart from secondary processes, the platelets were treated with indomethacin to prevent prostaglandin/thromboxane synthesis for all inducers; in addition, effects of secreted ADP, in the case of thrombin and A 23187, was prevented by the presence of creatine phosphate plus creatine phosphokinase ADP, epinephrine and A 23187, but not thrombin-induced primary aggregates, dispersed immediately upon addition of azide. Azide powerfully inhibited dense granule secretion induced by collagen, ADP and epinephrine as measured both by 14C-serotonin secretion and as judged by secondary aggregation. Shape change induced by ADP, thrombin or A 23187 was not affected. Azide had no effect on energy metabolism. Since the aggregation experiments were performed in the presence of indomethacin, and malondialdehyde formation from arachidonic acid was not affected by azide, it seemed unlikely that the inhibition by azide of platelet function was related to inhibition of synthesis of prostaglandins and thromboxanes. It is concluded that azide exerts its effects directly on the common pathway for platelet responses.

Mild haemostatic problems associated with congenital heterozygous alpha 2-antiplasmin deficiency.

A Dutch family, of which 13 members are heterozygotes, deficient for alpha 2-antiplasmin (alpha 2-AP) is reported. Clinical studies showed that 2 heterozygotes had a mild bleeding tendency, which presented as bleeding episodes after tooth extraction and after surgery and, in one patient, also as excessive menstruation. Laboratory investigations revealed an alpha 2-AP activity of 62% (51-71) (median and range) and an antigen level of 60% (60-66). The plasminogen binding as well as the fibrin binding properties of alpha 2-AP were normal. Plasminogen concentrations were significantly higher in the heterozygotes compared to the other family members. However, free plasminogen not bound to histidine-rich glycoprotein was not significantly different between these two groups. We propose that in this family the deficiency of alpha 2-AP is due to a decreased synthesis of a normal alpha 2-AP molecule. This present study brings the frequency of heterozygous alpha 2-AP deficient patients with a bleeding tendency to 13 out of 59 heterozygotes reported in the literature.

Phenotyping and genotyping of coagulation factor V Leiden.

The currently used activated Protein C resistance test demonstrated to be of limited diagnostic value for the detection of the mutant Factor V Leiden. Moreover, this assay is not useful for patients under anticoagulant therapy. A modification of the APC resistance test, applying Factor V deficient plasma is described which demonstrates a specificity and sensitivity of 1.0. The superiority of the modified APC resistance test over the existing APC resistance test was verified by genotyping. For that purpose, the Amplification Refractory Mutation System (ARMS) was applied to the detection of the G to A mutation at position 1691 in the gene encoding coagulation Factor V. The mutation at that position could be easily detected by using each of two allele-specific oligonucleotide primers concomitantly with one common primer in two separate polymerase chain reactions, thereby amplifying a fragment of 186 base-pairs of the Factor V gene.

Intravascular coagulation in liver transplantation--is it present or not? A comparison between orthotopic and heterotopic liver transplantation.

UNLABELLED: It is still not clear whether disseminated intravascular coagulation (DIC) contributes to the hemostatic disturbances in orthotopic liver transplantation (OLT). Theoretically the lack of hepatic clearance of procoagulant factors during the anhepatic period and the release of thromboplastic material from the graft might trigger DIC. During heterotopic liver transplantation (HLT) the host liver is left in situ and procoagulant factors may still be cleared; DIC, if present, may not occur until after reperfusion. The aim of the present study was to gain more insight into the underlying mechanism of the coagulation changes during liver transplantation by comparison of OLT and HLT. Thrombin-antithrombin-III complexes (TAT), and indicator of thrombin generation, fibrin degradation products (FbDP) and routine clotting times were assayed in 12 OLTs, 18 HLTs and in a control group of 10 partial hepatic resections (PHR). TAT increased dramatically after reperfusion to 136 micrograms/l in OLT and to 94 micrograms/l in HLT (p n.s.). In contrast, FbDP levels increased only in OLT, to a maximum of 13.8 micrograms/ml. Routine clotting times changed mildly and similarly in both OLT and HLT. CONCLUSIONS: Graft reperfusion triggers excessive thrombin formation, but there are no other signs of subsequent DIC. Any thrombin formed is probably rapidly inhibited by antithrombin-III. The rise in FbDP during OLT is the result of increased fibrinolysis, which occurred only in OLT and not in HLT.

Pharmacokinetics and tolerability of factor XIII concentrates prepared from human placenta or plasma: a crossover randomised study.

The pharmacokinetics and tolerability of factor XIII (FXIII) from plasma were compared with those of FXIII from placenta in a randomised, double-blind, crossover study involving 13 patients with congenital FXIII deficiency. Both FXIII activity and FXIII antigen were monitored. No difference was seen in the mean half-lives of the two preparations (9.3 days and 9.1 days for plasma and placenta FXIII activity, respectively). Response was similar for both preparations, but was slightly greater for FXIII from plasma (1.6 ormula: see text] vs 1.5 [formula: see text]). Similar results were found for recovery (65% vs 60%). The area under the data completed by extrapolation was significantly higher for FXIII from plasma. No differences between preparations in terms of efficacy or tolerability were observed. It can be concluded that treatment with FXIII concentrate from plasma is as efficient as with FXIII concentrate from placenta in terms of recovery and half-life. Both preparations were equivalent in terms of safety during the observation period. With the administration of monthly injections of approximately 30 U/kg serious bleeding events were prevented and no other serious adverse events occurred.

The molecular basis of antithrombin deficiency in Belgian and Dutch families.

The molecular basis of hereditary antithrombin (AT) deficiency has been investigated in ten Belgian and three Dutch unrelated kindreds. Eleven of these families had a quantitative or type I AT deficiency, with a history of major venous thromboembolic events in different affected members. In the other two families a qualitative or type II AT deficiency was occasionally diagnosed. DNA studies of the AT gene were performed, using polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) analysis, followed by direct sequencing of the seven exons and intron-exon junction regions. Six novel point mutations were identified: four missense, one nonsense mutation and a single nucleotide deletion near the reactive site, causing a frameshift with premature translation termination. In two kindreds the underlying genetic defect was caused by a whole gene deletion, known as a rare cause of AT deficiency. In these cases, Southern blot and polymorphism analysis of different parts of the AT gene proved useful for diagnosis. In another kindred a partial gene deletion spanning 698 basepairs could precisely be determined to a part of intron 3B and exon 4. In two type I and in both type II AT deficient families a previously reported mutation was identified. In all cases, the affected individuals were heterozygous for the genetic defect.

Stability and pharmacokinetics of flumazenil in the rat.

The pharmacokinetics of flumazenil in the rat were determined after 2.5 mg/kg intravenous and 25 mg/kg oral administration. Following intravenous administration flumazenil was rapidly eliminated with an extremely short terminal half-life (mean +/- SE, n = 8) of 8.3 +/- 0.3 min due to a large total blood clearance of 147 +/- 7 ml/kg/min combined with a relatively small volume of distribution at steady-state of 1.33 +/- 0.07 l/kg. After oral administration flumazenil was rapidly absorbed; however, the bioavailability was low (28 +/- 4%) and variable. Flumazenil was found to be unstable in rat blood in vitro and disappeared with a half-life (mean +/- SE, n = 5) of 8.3 +/- 1 min and 31 +/- 4 min at body and room temperature, respectively. The blood samples were stabilized by addition of sodium fluoride (NaF) and cooling to 0 degrees C. The samples had to be stored at -35 degrees C when analyzed at later times. Presumably esterases in rat blood are responsible for the observed instability. A sensitive HPLC assay to measure flumazenil concentrations in small blood samples is also described.

Involvement of the septohippocampal cholinergic system in representational memory.

To develop an animal model for testing muscarinic agonists, we examined the effects of cholinergic lesions with the ethylcholine aziridinium ion (AF64A) on two types of memory tasks. The tasks provided a distinction between representational and dispositional memory that could be measured in a single paradigm. Young, male Long-Evans rats were trained in a modified T-maze to learn both a discrimination task and a paired-run alternation task. Once animals learned the tasks, they were administered either saline or AF64A (5 nmol into each hippocampus) via stereotaxic technique. One week following surgery, saline-treated animals exhibited comparable performances (P greater than 0.2) on both the discrimination task (90.0 +/- 2.6% correct) and the alternation task (79.5 +/- 5.7%). In contrast, animals treated with AF64A showed a significant impairment of performance (P less than 0.005) on the alternation task (56.1 +/- 1.7%) as compared to the discrimination task (81.6 +/- 5.0%). Performance of the alternation task was significantly lower for AF64A-treated animals than for controls (P less than 0.02). AF64A-treated animals subsequently injected with pilocarpine (1.0 mg/kg, i.p.) showed moderate improvements in performance on the alternation task, while performance on the discrimination task remained unaffected. Immunocytochemical studies of choline acetyltransferase (ChAT) and tyrosine hydroxylase (TH) immunoreactivity indicated a loss of ChAT-positive cells in the septal region in AF64A-injected animals while TH-positive cells in the ventral tegmental area were unaffected by the treatment. The data suggest that AF64A can be used to produce selective lesions of the septohippocampal cholinergic system, which plays a greater role in representational memory than in dispositional memory.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochemical and behavioral responses of pilocarpine at muscarinic receptor subtypes in the CNS. Comparison with receptor binding and low-energy conformations.

Pilocarpine was tested biochemically in vitro for its ability to stimulate phosphoinositide (PI) turnover in the hippocampus (M1/M3 responses) where it displayed 35% of the maximal carbachol response with an EC50 value of 18 microM, and low-Km GTPase in the cortex (M2 response), where it had 50% of the maximal carbachol response with an EC50 value of 4.5 microM. Behaviorally, pilocarpine was able to restore deficits in a representational memory task (sensitive to M1 antagonists) produced by intrahippocampal injections of AF64A. Twenty-three low-energy conformations of protonated pilocarpine were generated using the program MacroModel. The data indicate that pilocarpine is a partial agonist at both M1 and M2 muscarinic receptors in the CNS. Behaviorally, with respect to the memory task, M1 effects of pilocarpine apparently predominate. It also is conceivable that different conformations of pilocarpine are active as agonists at different muscarinic receptor subtypes.

Evidence for a preferential involvement of M1 muscarinic receptors in representational memory.

The effects of intrahippocampal injections to the M1-selective antagonist pirenzepine and the M2-selective antagonist AF-DX 116 were examined on performance of a representational memory task in rats. Although both antagonists impaired performance, pirenzepine was more potent than AF-DX 116. Pirenzepine (70.8 +/- 2.8% correct) produced a greater deficit than AF-DX 116 (83.3 +/- 0.0%) at 70 micrograms, and the deficit at 10 micrograms (83.3 +/- 2.8%) was equal to that produced by 70 micrograms of AF-DX 116. The data provide additional support for the cholinergic hypothesis of memory and new information regarding the subtypes of muscarinic receptors likely to be involved in representational memory. Based on the greater susceptibility of representational memory to the effects of pirenzepine, it is suggested that M1 receptors in the hippocampus play a greater role in memory function than M2 receptors.

The reliability of Hickman catheter blood for the assessment of activation markers of coagulation and fibrinolysis in patients with hematological malignancies.

We studied the reliability of Hickman catheter (HC) blood for the determination of activation markers of coagulation and fibrinolysis in 14 patients with hematological malignancies. 10 of 22 HC samples were contaminated with heparin, probably as a consequence of our institutional antithrombotic catheter care, and withdrawn from statistical analysis. Prothrombin activation fragment F1.2, thrombin-antithrombin III complexes, soluble fibrin, degradation products of fibrinogen and fibrin, total degradation products, alpha 2-antiplasmin and antithrombin III levels withdrawn from Hickman catheters did not differ significantly from peripheral venous blood samples. Fibrinogen levels were slightly but significantly higher in peripheral venous blood samples. We conclude that Hickman catheter blood gives reliable results for the determination of activation markers of coagulation and fibrinolysis in patients with hematological malignancies, but may be less useful in clinical practice due to frequent heparin contamination.

Pharmacokinetics, enantiomer interconversion, and metabolism of R-apomorphine in patients with idiopathic Parkinson's disease.

The pharmacokinetics and metabolism of R-apomorphine were determined in 10 patients with idiopathic Parkinson's disease after intravenous infusion of 30 micrograms.kg-1 in 15 min. Specifically, emphasis was on enantiomeric interconversion into S-apomorphine and on the formation of apocodeine and isoapocodeine, since these metabolites may interfere with the pharmacodynamics of R-apomorphine. The pharmacokinetics of R-apomorphine in plasma were determined using an enantioselective high-performance liquid chromatography assay. In most patients, the plasma concentration versus time profile was characterized by a biexponential function. The values of relevant pharmacokinetic parameters were as follows: clearance 40 +/- 15 ml.min-1.kg-1, volume of distribution at steady state 1.6 +/- 0.5 l.kg-1, and terminal half-life 41 +/- 13 min. No measurable concentrations of S-apomorphine were detected in plasma, indicating that enantiomeric interconversion does not occur in vivo. Furthermore, no measurable concentrations of the methylated metabolites apocodeine and isoapocodeine could be detected in plasma. The metabolism of apomorphine was characterized on basis of the excretion of unchanged R-apomorphine, S-apomorphine, apocodeine, isoapocodeine, and their respective sulfate and glucuronide conjugates in urine. The total excretion of unconjugated S-apomorphine, apocodeine, and isoapocodeine was less than 0.1% of the administered dose. The total excretion of unchanged apomorphine, apomorphine sulfate, and apomorphine glucuronide amounted to 0.3 +/- 0.4%, 3.8 +/- 1% and 6.0 +/- 2.2% of the administered dose, respectively. The findings of this study show that on intravenous administration, S-apomorphine and the metabolites apocodeine and isoapocodeine are unlikely to interfere with the pharmacologic actions of R-apomorphine in patients with idiopathic Parkinson's disease. Furthermore, no pharmacokinetic interaction between R-apomorphine and catechol-O-methyl transferase inhibitors is expected.

Heparin-induced thrombocytopenia and thrombosis: a potential fatal complication in a routine treatment.

Heparin-induced thrombocytopenia (HIT) is a severe complication of heparin therapy. Life-threatening thromboembolism (HITT) may occur in a large number of patients with HIT. In this article diagnostic problems and the clinical course of 4 typical patients are described. Diagnosis was based on the occurrence of thrombocytopenia during heparin therapy and was confirmed in vitro by an ELISA to heparin-platelet factor 4 antibodies, heparin-induced platelet activation assay (HIPAA) or the platelet aggregation assay (PAA). Thrombotic complications developed in 2 patients, one of whom suffered a fatal embolism after accidentally rechallenging with low-dose heparin which was used to maintain the patency of an intravascular catheter. After discontinuation of heparin the thrombocyte count rapidly increased to normal values during treatment with the heparinoid danaparoid (Orgaran) without complications.

Selection of asymptomatic HIV carriers for antiviral therapy.

In order to find suitable markers for selection and monitoring of antiviral therapy in asymptomatic HIV-infected patients, we evaluated 18 anti-HIV positive individuals at three monthly intervals by HIV culture, HIV antigen, and core (p24) antibody testing as well as by measurement of lymphocyte subsets. Consistent results were obtained with HIV antigen, p24 antibody testing and T4 cell enumeration, whereas results of virus detection were variable. Therefore cumbersome and expensive virus culture is not of use in selecting patients for antiviral therapy. On the basis of our results and recent literature we currently propose using absence of p24 antibodies, presence of HIV antigen and low or falling T4 cells as eligibility criteria for antiviral therapy in asymptomatic infected individuals.

Prophylaxis of thrombosis in antithrombin III-deficient women during pregnancy and delivery.

Prophylactic treatment of two asymptomatic antithrombin III (AT III) -deficient women with oral anticoagulants and heparin during pregnancy and after delivery appeared to be feasible. Oral anticoagulants caused no change in plasma AT III levels in either woman. A high dose of heparin intravenously in one of them caused a strong reduction in the AT III level which was corrected by infusion of AT III concentrate during delivery. A low dose of heparin subcutaneously in the other caused no significant change in plasma AT III levels. There is probably no need for AT III substitution during an uncomplicated pregnancy.

Fatal ischaemic brain oedema after early thrombolysis with tissue plasminogen activator in acute stroke.

Two patients with acute major, disabling cerebral infarction with presumed middle cerebral artery occlusion were treated with the clot specific thrombolytic agent tissue plasminogen activator roughly three and a half hours after the onset of symptoms. Both patients had a normal computed tomography (CT) scan before treatment. No appreciable systemic bleeding complications occurred, apart from bruising. One patient had bleeding into the subarachnoid space from a microscopic angioma, which was found at necropsy. Haematological monitoring of the two patients showed pronounced fibrinogenolysis and alpha 2 antiplasmin consumption in one. One patient showed transient improvement during the infusion. In both cases extensive infarction, partly haemorrhagic in one, with massive concomitant oedema was found on repeated CT. Both patients deteriorated and eventually died as a consequence of transtentorial herniation. In the one patient who came to necropsy a moderate, probably pre-existing smooth stenosis of the ipsilateral carotid artery was found, all cerebral vessels being patent. It is concluded that thrombolytic treatment with a clot specific agent such as tissue plasminogen activator started three to four hours after a major ischaemic stroke may be hazardous, not because of haemorrhagic transformation of the original ischaemia but because early reperfusion may promote massive, potentially fatal cerebral oedema.

[Possibilities and limitations of heterotopic auxiliary liver transplantation]. / Möglichkeiten und Grenzen der heterotopen auxiliären Lebertransplantation.

Although auxiliary heterotopic liver transplantation offers theoretical advantages over orthotopic liver replacement, clinical results have heretofore been dismal. After development of a technique of reduced size liver grafts provided with portal and arterial blood and venous drainage via the suprahepatic V. cava (HLT) in experimental animals, this method was applied in 21 transplantations in 19 patients. 11 of 16 patients with chronic liver insufficiency and one of three patients with fulminant liver failure survived transplantation for at least 1 year. HLT was well tolerated even by high-risk patients. Possibilities and limitations of this novel approach are discussed.