Pipecolic acid induces oxidative stress in vitro in cerebral cortex of young rats and the protective role of lipoic acid.

Pipecolic acid (PA) levels are increased in severe metabolic disorders of the central nervous system such as Zellweger syndrome, infantile Refsum disease, neonatal adrenoleukodystrophy and hyperlysinemia. The affected individuals present progressive neurological dysfunction, hypotonia and growth retardation. The mechanisms of brain damage of these disorders remain poorly understood. Since PA catabolism can produce H2O2 by oxidases, oxidative stress may be a possible mechanism involved in the pathophysiology of these diseases. Lipoic acid (LA) is considered an efficient antioxidant and has been shown to prevent oxidative stress in experimental models of many disorders of the neurologic system. Considering that to our knowledge no study investigated the role of PA on oxidative stress, in the present work we investigated the in vitro effects of PA on some oxidative stress parameters and evaluated the LA efficacy against possible pro-oxidant effects of PA in cerebral cortex of 14-day-old rats. The activities of catalase (CAT), glutathione peroxidase (GPx), glucose 6-phosphate dehydrogenase (G6PD), and glutathione S-transferase (GST) along with reduced glutathione (GSH) content were significantly decreased, while superoxide dismutase (SOD) activity and thiobarbituric acid-reactive substances (TBA-RS) were significantly enhanced by PA. LA was able to prevent these effects by improving the activity of antioxidant enzymes, increasing GSH content and reducing TBA-RS. In contrast, glutathione reductase and 6-phosphogluconate dehydrogenase activities and sulfhydryl content were not affected. Taken together, it may be presumed that PA in vitro elicits oxidative stress and LA is able to prevent these effects.

Photopic electroretinograms of mGluR6-deficient mice.

PURPOSE: To study the properties of the photopic electroretinograms (ERGs) of the metabotropic glutamate receptor subtype 6 (mGluR6)-deficient mice and to investigate the contribution of cone ON-and OFF-pathways to the mouse photopic ERGs. METHODS: Photopic ERGs were recorded from mGluR6-deficient and wild-type mice. Photopic ERGs were also recorded after an intravitreous injection of cis-2,3 piperidine dicarboxylic acid (PDA) to block the transmission of signals from the photoreceptors to the OFF-bipolar cells, horizontal cells, and other inner retinal neurons. RESULTS: The amplitude of the b-wave of the photopic ERG was severely reduced in mGluR6-deficient mice, but a small, slow, positive component was seen after the a-wave. Intravitreous injection of PDA eliminated this positive component. CONCLUSIONS: The mGluR6-deficient mouse is a useful animal model to study the contribution of the ON-and OFF-pathways to the mouse ERG.

Multifocal electroretinogram in rhodopsin P347L transgenic pigs.

PURPOSE: Neural ectopic rewiring in retinal degeneration such as retinitis pigmentosa (RP) may form functional synapses between cones and rod bipolar cells that cause atypical signal processing. In this study, the multifocal electroretinograms (mfERGs) of a large animal model of RP, the rhodopsin P347L transgenic (Tg) pig, were measured to examine the sources and nature of altered signal processing. METHODS: mfERG responses from a 6-week-old Tg pig were recorded before and after sequential application of tetrodotoxin (TTX), N-methyl-D-aspartate (NMDA), 2-amino-4-phosphonobutyric acid (APB), and cis-2,3-piperidinedicarboylic acid (PDA), to identify contributions to the retinal signal from inner retinal neurons, the ON-pathway, the OFF-pathway, and photoreceptors. The mfERG response contributions from different retinal components of in the Tg eyes were estimated and compared with control data from eyes of age-matched wild-type (WT) pigs. RESULTS: There was a prominent difference in the estimates of the inner retinal response and ON-bipolar cell pathway contribution between the Tg and WT mfERG responses. In particular, the early components of the inner retinal contribution were obviously altered in the Tg mfERG. The inner retinal components at approximately 24 and 40 ms appeared to be inverted. Differences in the estimates of OFF-bipolar cell pathway contributions were minimal. There was no change in cone cell responses in the Tg mfERG. CONCLUSIONS: In Tg retinas, ectopic synapses formed between cones and rod bipolar cells probably altered signal processing of the ON-bipolar cell pathway. In response to the altered visual signal input from the outer retina, signal processing in inner retinal neurons was also modified.

Transitioning from argatroban to warfarin in heparin-induced thrombocytopenia: an analysis of outcomes in patients with elevated international normalized ratio (INR).

BACKGROUND: Heparin-induced thrombocytopenia (HIT) can lead to catastrophic thromboembolic complications and requires treatment with an alternative, rapidly active anticoagulant, such as a direct thrombin inhibitor (DTI), either to prevent or treat these complications. Switching to oral warfarin after initial treatment with a DTI is necessary in most patients. Most references related to warfarin suggest that an increased risk for bleeding will occur with elevated international normalized ratios (INRs) > 4.6. In patients receiving argatroban, it is not uncommon to achieve an INR > 4 during this transition. Because the clinical outcomes in patients achieving an INR > 4 during combined argatroban/warfarin therapies for HIT are not well described, we evaluated the clinical outcomes of 111 patients with this phenomenon. METHODS: We identified patients from the prospective studies of argatroban anticoagulation, Argatroban-911 and Argatroban-915. Data collected from these studies included death from all causes, amputation, new thrombosis, major bleeding, INR values, argatroban doses, aPTT values, platelet counts, and duration of therapy. RESULTS: Patients were on argatroban monotherapy for a median of 2.8 (0.1-8.1) days, and on cotherapy for a median of 3.7 (0.9-12.8) days. The median platelet count was 70.9 (18-325) x 10(9)/L at the time of HIT diagnosis and increased to 94 (30-324) x 10(9)/L by the time warfarin was initiated. At a median argatroban dose of 1.4 (0.2-2.0) mcg/kg/min, the maximum INR ranged from 4.1 to 21.2 (median 6.4, n = 111) and the corresponding aPTT ranged from 48.1 to 105 (median 71, n = 93) seconds. After argatroban cessation, the first recorded INR within 4 to 24 hours ranged from 1.5 to 12.5 (median 2.9, n = 58). Adverse clinical outcomes occurred in 9 (8.1%) patients during cotherapy and in 12 (10.8%) patients after argatroban anticoagulation was discontinued. Adverse clinical outcomes included 7 cases of new thrombosis, 3 amputations, 12 deaths and 1 major bleed. Eleven of 12 (91.7%) patients died due to causes other than thrombosis, and most deaths (83%) occurred following cotherapy. Five (4.5%) patients developed new thrombosis during argatroban/warfarin cotherapy despite an INR > 4. In contrast only 1 (0.9%) patient experienced major bleeding. CONCLUSION: In patients receiving argatroban/warfarin cotherapy and with an elevated INR > 4, the risk for thrombosis exceeds the risk of bleeding. Traditional paradigms concerning elevated INRs and warfarin may need to be redesigned for the patient population on cotherapy with direct thrombin inhibitors.Abbreviated Abstract. The clinical outcomes of 111 patients with INRs > 4 while on combined argatroban (dose < or = 2 mcg/kg/min) and warfarin were evaluated. Adverse clinical outcomes (7 new thrombosis, 3 amputations, 12 deaths and 1 major bleed) occurred in 21 patients. Eleven deaths were due to causes other than thrombosis. Five patients developed new thrombosis while only 1 had major bleeding. The risk for thrombosis exceeds the risk of bleeding in patients with HIT despite an INR > 4.

Interpreting the International Normalized Ratio (INR) in individuals receiving argatroban and warfarin.

The effects of argatroban, a direct thrombin inhibitor, on the International Normalized Ratio (INR), activated partial thromboplastin time (aPTT) and functional factor X during warfarin co-administration were established to provide means to interpret INRs during argatroban/warfarin co-therapy. Twenty-four subjects receiving warfarin (7.5 mg, day 1; 3-6 mg/day, days 2-10) and argatroban (1-4 microg/kg/min over 5 h, days 1-11) were assessed daily for these coagulation parameters prior to argatroban infusion (warfarin "monotherapy") and at its conclusion ("co-therapy"). Argatroban increased aPTTs dose-dependently. Co-therapy INR increased linearly with monotherapy INR, with slope sensitive to argatroban dose and thromboplastin used. Prediction errors for monotherapy INRs were < or =+/- 0.4 for argatroban 1-2 microg/kg/min but > or = +/-1.0 for higher doses. Despite co-therapy INRs >7, no major bleeding occurred. Factor X remained > or =37% of normal. Therefore, the predictable effect of argatroban (< or =2 microg/kg/min only) [corrected] on INRs during warfarin co-therapy allows for reliable prediction of the level of oral anticoagulation.

Pharmacological effects of a novel recombinant hirudin, CX-397, in vivo and in vitro: comparison with recombinant hirudin variant-1, heparin, and argatroban.

The novel recombinant hirudin analog CX-397 was investigated with respect to its pharmacological activity and antithrombin profiles in vivo and in vitro. In three different types of thrombosis models in rats, including stasis and thrombin-induced venous, glass surface-activated arterio-venous shunt, and ferric chloride-induced arterial thrombosis models, CX-397 and rHV-1 elicited potent antithrombotic effects, where the minimum effective doses of rHV-1 tended to be higher than those of CX-397 in the arterio-venous shunt and arterial thrombosis models. The hemorrhagic risk of CX-397 in template bleeding in rats was not higher than that of rHV-1, indicating that CX-397 is superior to rHV-1 for treating the platelet-dominant type of thrombosis. However, no differences were detected between CX-397 and rHV-1 in their effects on in vitro coagulation times and thrombin-induced platelet aggregation, suggesting the possibility that some unknown mechanisms other than simple thrombin inhibition are also involved in their antithrombotic actions.

Effect of a synthetic factor Xa inhibitor, YM-60828, on blood vessel patency in combination with a thrombolytic agent and on blood loss from the operation site in a rat model of arterial thrombosis.

We examined the adjunctive effect of a novel factor Xa inhibitor, YM-60828, on vessel patency and blood loss from the operation site after successful thrombolysis with a modified tissue-type plasminogen activator (moPA) in an electrically-induced carotid artery thrombosis model in rats. Five minutes after the induction of occlusive thrombus, a test drug (YM-60828, argatroban, heparin or saline) was administered by i.v. bolus injection followed by continuous infusion. Thrombolysis was induced with moPA by i.v. bolus injection at a dose of 650,000 IU/ kg. YM-60828 at 1 mg/kg i.v. followed by 3 mg/kg/h significantly prevented reocclusion, increased the duration of patency, and improved vessel patency after successful thrombolysis without any significant increase in blood loss from the operation site. Argatroban at 1 mg/kg i.v. followed by 3 mg/kg/h and heparin at 300 U/kg i.v. followed by 150 U/kg/h also significantly improved these parameters, but were accompanied by a significant increase in blood loss. These results suggest that the factor Xa inhibitor YM-60828 may be a potent and useful adjunctive agent with a lower risk of bleeding complications than argatroban and heparin in thrombolytic therapy.

Acute toxicity of bupivacaine metabolites in mice.

1. This study was designed to document the acute toxicity of two metabolites of bupivacaine, desbutylbupivacaine (2,6, desbutylbupivacaine; PPX) and pipecolic acid in mice. All the compounds were administered by the intraperitoneal (i.p.) route. 2. The mean convulsant activity was 100% for controls, 30, 100, 100, 100 and 90% for 400, 200, 150, 125 and 112.5 mg/kg i.p. of PPX, respectively, and 0% for the animals receiving pipecolic acid. 3. The acute induced mortality was 60% for bupivacaine control group (50 mg/kg/i.p.), 60, 30 and 0% for 800, 400 and 200 mg/kg of pipecolic acid, respectively, and 100, 90, 60, 80 and 10% for 400, 200, 150, 125 and 112.5 mg/kg i.p. of PPX, respectively. 4. The time to convulse was 158 +/- 16 s for bupivacaine, 230 +/- 30, 270 +/- 24, 255 +/- 21, 442 +/- 84 and 418 +/- 32 s for 200, 150, 125, 112.5 and 100 mg/kg i.p. of PPX, respectively; any pipecolic acid treated animal have convulsed. 5. In conclusion, the present study demonstrated that PPX is more toxic than expected since we found that its induced mortality was approximately three times that found for bupivacaine and its CNS toxicity was about two times that of bupivacaine.

[Hydrochlorides of N-hydroxyalkyl-alpha-piperidine carboxylic arylamides possessing antiarrhythmic activity]. / Khlorgidraty nekotorykh arilamidov N-oksialkil-al'fa-piperidinkarbonovykh kislot, obladaiushchie antiaritmicheskoi aktivnost'iu.

It has been demonstrated in experimental arrhythmias of animals (aconitine one in rats, strophanthine in guinea-pigs, creation of an "ectopic focus of excitation" in cats by electric stimulation) that new compounds, the derivatives of piperidine carboxylic acids, possess antiarrhythmic activity. In doses of 6-23 mg/kg, the compounds under study prevented the development of the mixed type arrhythmia induced by intravenous injection of aconitine, prolonged the time of survival and prevented the animals' death during intravenous injection of the arrhythmogenic doses of strophanthine, prolonged the refractory period, and increased the threshold of ventricular fibrillation in cats. As regards the power of the antiarrhythmic effect, the compounds under study--hydrochlorides of arylamides of N-hydroxyalkly-alpha-piperidine carboxylic acids--approach lidocaine, yield to marcaine, and exhibit a broader range of therapeutic action.

Anticonvulsant and proconvulsant properties of a series of structural isomers of piperidine dicarboxylic acid.

Anticonvulsant and convulsant effects of various piperidine dicarboxylic acids have been evaluated following their intracerebroventricular (i.c.v.) or intraperitoneal (i.p.) injection in DBA/2 mice, a strain of mice genetically susceptible to sound-induced seizures. Protection against sound-induced seizures occurred after intraventricular administration of (+/-)cis-2,3-piperidine dicarboxylic acid (0.017-0.045 mumol), (+/-)trans-2,3-piperidine dicarboxylic acid (0.018-0.33 mumol) and (+/-)cis-2,4-piperidine dicarboxylic acid (0.57-1.68 mumol). Protection against sound-induced seizures occurred after intraperitoneal injection of (+/-)cis-2,3-piperidine dicarboxylic acid (0.52-1.8 mmol/kg). Myoclonus or convulsions occurred at various times after the intraventricular injection of cis-2,3-piperidine dicarboxylic acid, trans-2,3-, cis-2,4-, cis-2,5- and cis-2,6-, piperidine dicarboxylic acids, and after the intraperitoneal injection of trans-2,3-piperidine dicarboxylic acid. The latter effect was blocked by pretreatment with 2-amino-7-phosphonoheptanoic acid (0.33 mmol/kg, i.p.) a potent and specific antagonist of excitation induced by N-methyl-D-aspartate. The anticonvulsant action of cis-2,3-piperidine dicarboxylic acid and the convulsant action of trans-2,3-piperidine dicarboxylic acid were associated with predominant antagonist and agonist actions respectively, at receptors preferring N-methyl-D-aspartate.

On the excitotoxic properties of quinolinic acid, 2,3-piperidine dicarboxylic acids and structurally related compounds.

To obtain information about the receptors which mediate the neurotoxic actions of quinolinic acid, a series of pyridine dicarboxylates and piperidine dicarboxylates and structurally related compounds were tested for their neurotoxic effects following intrastriatal or intrahippocampal infusion in the rat, and for their activity in assays of binding and uptake sites for acidic amino acids. Of the compounds tested, only cis- and trans-2,3-piperidine dicarboxylates and quinolinic acid showed pronounced neurotoxic effects. At 600 nmol, 2,6- and 3,4-pyridine dicarboxylates were weakly active and the remaining compounds were inactive in both brain regions. After injection into the striatum of the adult rat, trans-2,3-piperidine dicarboxylate, quinolinic acid and cis-2,3-piperidine dicarboxylate caused axon-sparing neuronal degeneration as assessed by light microscopic and neurochemical methods, the threshold doses being 12, 24 and 120 nmol, respectively. In the striatum of the 7-day old rat, 30 nmol quinolinic acid or 600 nmol cis-2,3-piperidine dicarboxylate were inactive. Small doses of cis-2,3-piperidine dicarboxylate (60 nmol) and quinolinic acid (30 nmol) injected into the adult rat hippocampus resulted in a preferential loss of pyramidal neurons. In larger doses granule cells also degenerated. In contrast, trans-2,3-piperidine dicarboxylate was equally toxic to hippocampal neurons, regardless of the dose used. No "distant" neuronal damage was observed after the intracerebral application of any test compound. Equimolar amounts of (-)-2-amino-7-phosphonoheptanoic acid completely blocked the neurotoxic effects of quinolinic acid, cis- and trans-2,3-piperidine dicarboxylate after injection into the striatum or hippocampus. None of the analogs tested were good inhibitors of Cl--dependent or independent binding of L-[3H]glutamate, [3H]kainate or high-affinity, Na+-dependent uptake of L-glutamate in striatal or hippocampal tissue at 1 mM. The results indicate that the receptors mediating the neurotoxic effects of these compounds have strict structural requirements for activation. Whereas the excitotoxic characteristics of trans-2,3-piperidine dicarboxylate suggest a direct action on N-methyl-D-aspartate receptors, the properties of quinolinic acid and cis-2,3-piperidine dicarboxylate are far more complex and make categorization of their receptor-interactions difficult. Indirect mechanisms may account for the excitotoxicity of quinolinic acid and cis-2,3-piperidine dicarboxylate.