The effects of perinatal hypoxia on pentylenetetrazol-induced seizures in developing rats

The effects of perinatal hypoxia on susceptibility to seizures due to a single dose (55 mg/kg, i.p.) of pentylenetetrazol (PTZ) were examined, in 15-, 20-, 30-, 60- and 90 to 120-day-old rats. The rats exposed to hypoxia at 10 days of age and the unexposed controls showed similar developmental changes in the types of seizures, ictal electro-encephalograms and severity scores, the last being lowest at 30 days of age. However, the susceptibility to seizures induced by PTZ, which was measured by the mean number of generalized convulsions (GCs) as well as a mean duration of the 1st GC and severity score, was more enhanced in the rats exposed to perinatal hypoxia than in the control group at every age. The present study suggests that hypoxia in rats results in greater seizure susceptibility throughout the developmental period.     

Asymmetric syntheses of pipecolic acid and derivatives

Summary Results in the field of asymmetric synthesis of pipecolic acid derivatives are reviewed. Three sections describe the asymmetric syntheses of the title compounds (i) from the chiral pool (-amino acids or carbohydrates) (ii) using a chiral auxiliary either derived from terpenes,-amino acids, tartaric acid, an amine or-amino alcohols (iii) by means of asymmetric catalysis.     

Uptake of piperidine and pipecolic acid by synaptosomes from mouse brain

Piperidine is actively transported into the synaptosomal fraction of adult mouse brain. The transport mechanism appears to be Na⁺ independent but is temperature dependent and sensitive to ouabain. Analysis of kinetic experiments indicates only a low-affinity transport system to be present. By contrast the uptake ofD,L-[³H]pipecolic acid at a concentration of 4×10^–7 M was temperature and Na⁺ dependent, ouabain sensitive, and revealed a two-component system with aK _m =3.9±0.17×10^–6 M,V _max=129±6 pmol/mg protein/3 min for the high-affinity system and aK _m =90.2±4.3×10^–6 M,V _max=2.45±0.19 nmol/mg protein/3 min for the low-affinity system. Compounds structurally related to pipecolic acid such as glycine,l-proline, 4-amino-n-butyric acid, and 5-amino-n-valeric acid showed an inhibitory effect on uptake at a concentration of 10^–4 M. The demonstration of biosynthesis of pipecolic acid in mouse brain and the presence of a high-affinity sodium-dependent uptake system suggest a physiological role of this substance in the central nervous system.     

Modulation of retinoic acid receptor alpha activity by lysine methylation in the DNA binding domain

Metabolic labeling and detection with a methylated lysine-specific antibody confirm lysine methylation of RAR alpha in mammalian cells. We previously reported Lys (347) trimethylation of mouse retinoic acid receptor alpha (RAR alpha) in the ligand binding domain (LBD) that affected ligand sensitivity of the dissected LBD. Here we report two monomethylated residues, Lys (109) and Lys (171) identified by LC-ESI-MS/MS in the DNA binding domain (DBD) and the hinge region, which affect retinoic acid (RA) sensitivity, coregulator interaction and heterodimerization with retinoid X receptor (RXR) in the context of the full-length protein. Constitutive negative mutation at Lys (109), but not Lys (171), reduces RA-dependent activation. Methylation at Lys (109) plays a more dominant role than trimethylation at Lys (347) in terms of RA activation of the full-length receptor. Lys (109) is located in a homologous sequence (CEGC K GFFRRS) of the DBD in RARs and is conserved in the nuclear receptor superfamily even across the species boundary. This study uncovers a potential role for monomethylation at Lys (109) in coordinating the synergy between DBD and LBD for ligand-dependent activation of RAR alpha.     

The conversion of lysine into piperidine,cadaverine, and pipecolic acid in the brain and other organs of the mouse

The biosynthesis of piperidine, a possible neuromodulator, and its presumed precursors cadaverine and pipecolic acid, has been investigated in the mouse under in vitro conditions. Conversion of lysine into piperidine was observed only in the intestines and is probably caused by the intestinal flora. Formation of cadaverine and pipecolic acid from lysine was observed in the brain, liver, kidney, and large intestine. In addition, pipecolic acid was formed in the heart. The possible contributions of the diet and of the intestinal bacteria to the endogenous pool(s) of piperidine are discussed.     

Antioxidant activity elicited by low dose of caffeine attenuates pentylenetetrazol-induced seizures and oxidative damage in rats

Although caffeine supplementation has a beneficial effect on people with neurological disorders, its implications for oxidative damage related to seizures are not well documented. Thus the aim of this study was to investigate the effects of two weeks caffeine supplementation (6 mg/kg; p.o.) on seizures and neurochemical alterations induced by pentylenetetrazol (PTZ 60 mg/kg i.p.). Statistical analyses showed that long-term rather than single dose caffeine administration decreased the duration of PTZ-induced seizures in adult male Wistar rats as recorded by cortical electroencephalographic (EEG) and behavioral analysis. The quantification of EEG recordings also revealed that caffeine supplementation protected against a wave increase induced by PTZ. Neurochemical analyses revealed that caffeine supplementation increased glutathione (GSH) content per se and protected against the increase in the levels of thiobarbituric acid reactive substances (TBARS) and oxidized diclorofluoresceine diacetate (DCFH-DA). Also, caffeine prevent the decrease in GSH content and Na⁺, K⁺-ATPase activity induced by PTZ. Our data also showed that the infusion of L-buthionine sulfoximine (BSO; 3.2 μmol/site i.c.v), an inhibitor of GSH synthesis, two days before injecting PTZ reversed the anticonvulsant effect caused by caffeine. BSO infusion also decreased GSH content and Na⁺, K⁺-ATPase activity. However, it increased DCFH-DA oxidation and TBARS per se and reversed the protective effect of caffeine. Results presented in this paper support the neuroprotective effects of low long-term caffeine exposure to epileptic damage and suggest that the increase in the cerebral GSH content caused by caffeine supplementation may provide a new therapeutic approach to the control of seizure.     

Sex differences in modulating blood brain barrier permeability by NO in pentylenetetrazol-induced epileptic seizures

Susceptibility to epilepsy as well as BBB dysfunction in some pathological conditions varies depending on sex difference. It has recently been shown that systemically given NO donor and antagonists modify the nature of seizures induced by PTZ (pentylenetetrazol) differently in male and female rats. This study investigates the role of NO on BBB permeability in PTZ seizures with sex differences using NO donor, sodium nitroprusside (SNP), and NOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME). Nitrite+nitrate levels as indices of NO generation in the brain were also assessed. L-NAME prolonged seizure latency in male rats, seizure intensity and seizure duration were lessened. L-NAME depicted opposite effects in seizure nature in female rats. SNP prolonged seizure latency, while seizure intensity and duration were lessened only in female rats. L-NAME in male rats increased L-NAME use in female rats (not in male rats) which resulted in a more leaky BBB especially in midbrain, thalamus, hippocampus, corpus striatum and cerebellum whereas SNP use in male rats and not in female rats resulted in pronounced BBB opening in all brain regions studied than PTZ per se. L-NAME while decreasing nitrite+nitrate levels in male rat brains, acted in an opposite fashion in females. SNP use depicted an inverse picture with respect to L-NAME, with an opposite action in different sexes. This study reveals that NO effect on BBB in PTZ-induced seizures depends unequivocally on sex difference. The sex-dependent action of NO in seizures and in CNS pathologies warrants further investigation.     

Acute creatine administration improves mitochondrial membrane potential and protects against pentylenetetrazol-induced seizures

A growing body of evidence indicates that creatine (Cr) exerts beneficial effects on a variety of pathologies where energy metabolism and oxidative stress play an etiological role. However, the benefits of Cr treatment for epileptics are still shrouded in controversy. In the present study, we found that acute Cr treatment (300 mg/kg, p.o.) prevented the increase in electroencephalographic wave amplitude typically elicited by PTZ (30, 45 or 60 mg/kg, i.p.). Cr treatment also increased the latency periods of first myoclonic jerks, lengthened the latency periods of the generalized tonic–clonic seizures and reduced the time spent in the generalized tonic–clonic seizures induced by PTZ (60 mg/kg). Administration of PTZ (all doses) decreased Na⁺, K⁺-ATPase activity as well as adenosine triphosphate (ATP) and adenosine diphosphate levels in the cerebral cortex, but Cr treatment prevented these effects. Cr administration also prevented increases in xanthine oxidase activity, adenosine monophosphate levels, adenosine levels, inosine levels and uric acid levels that normally occur after PTZ treatment (60 mg/kg, i.p.). We also showed that Cr treatment increased the total Cr (Cr + PCr) content, creatine kinase activity and the mitochondrial membrane potential (ΔΨ) in the cerebral cortex. In addition, Cr prevented PTZ-induced mitochondrial dysfunction characterized by decreasing ΔΨ, increasing thiobarbituric acid-reactive substance levels and increasing protein carbonylation. These experimental findings reinforce the idea that mitochondrial dysfunction plays a critical role in models of epileptic seizures and suggest that buffering brain energy levels through Cr treatment may be a promising therapeutic approach for the treatment of this neurological disease.     

Recent advances in asymmetric synthesis of pipecolic acid and derivatives

Summary. This review covers the literature relating to asymmetric syntheses of pipecolic acid derivatives from 1997 to present. This review is organized according to the position and the degree of substitution of the piperidinic cycle. In a first section, syntheses of pipecolic acid itself are described. Then, successively, syntheses of C-3, C-4, C-5, C-6 substituted pipecolic acid derivatives are reported. Finally, syntheses of unsaturated pipecolic acid derivatives are presented before the last part devoted to the polysubstituted pipecolic acid derivatives.     

The oxidation of pipecolic acid in preincubated soils

Chernozem soil was preincubated with 0.1% glucose, glucose plus ammonium nitrate, hydrolyzed casein or amino acids for five days and with 1% wheat straw, pectin, peptone or cellulose for ten days. The soil was modified by this treatment so that the pipecolic acid was oxidized in two steps, as was shown by two peaks on the graphic plot of the oxygen uptake. In the control soil sample preincubated with water, the oxygen consumption curve formed one peak. In brown soil it was necessary to increase the concentration of glucose and hydrolyzed casein to 1% to obtain the oxidation of pipecolic acid in two phasos. Approximately a third of the oxygen for the complete oxidation of pipecolic acid was consumed in the first phase of oxidation, another third in the second phase of oxidation. A relation was found between the occurrence of the second period of oxidation of pipecolic acid and the amount of organic carbon in the soil.     

Protective effect of safranal on pentylenetetrazol-induced seizures in the rat: Involvement of GABAergic and opioids systems

The aim of the present study was to evaluate the effects of safranal, an active constituent of Crocus sativus L. stigmas, on seizures induced by pentylenetetrazol. Intracerebroventricular (i.c.v.) microinjection of safranal (4.84, 9.68 and 24.2 micromol) had no effects on tonic and clonic phases as well as mortality upon seizures induced by PTZ (90mg/kg body wt., i.p.). Peripheral administration of safranal (72.75, 145.5 and 291 mg/kg body wt., i.p.), however, induced a dose-dependent decrease in the incidence of both minimal clonic seizures (MCS) (145.5 mg/kg body wt., p<0.01) and generalized tonic-clonic seizures (GTCS) (145.5 mg/kg body wt., p<0.001) following PTZ administration. Safranal also increased MCS and GTCS latency, significantly. Percent of protection against GTCS was 30%, 100% and 100% and mortality protection percent was 40%, 100% and 100% for the mentioned doses, respectively. Pretreatment with flumazenil (5 nmol, i.c.v.) and naloxone (5.5 nmol, i.c.v. and 2 mg/kg body wt., i.p.), 15 min prior to safranal administration (145.5 mg/kg body wt., i.p.), abolished the protective effect of safranal on MCS. Flumazenil also decreased the effect of safranal on incidence as well as latency of GTCS, significantly. These effects were not, however, significant for naloxone (5.5 nmol, i.c.v. and 2mg/kg body wt., i.p.). Results of this study demonstrated that safranal could exert anticonvulsant activity in the PTZ model and this effect may be mediated, at least partly, through GABA(A)-benzodiazepine receptor complex.     

Serotonin mediation of the protective effect of clonidine against pentylenetetrazol-induced seizures in rats

An intraperitoneal injection of 0.5 mg/kg clonidine significantly increased the latency to the first convulsion and reduced tonic seizures and mortality caused by pentylenetetrazol (PTZ), 90 mg/kg, administered subcutaneously to rats. 1 mg/kg clonidine produced similar effects except that tonic seizures were not significantly affected. No effect was observed with 0.01 or 0.1 mg/kg clonidine. Metergoline (1 mg/kg) and methysergide (10 mg/kg), administered intraperitoneally, completely prevented the effect of 0.5 mg/kg clonidine on PTZ-induced seizures. An intraperitoneal injection of 5 mg/kg of d-fenfluramine, a releaser of 5HT from nerve terminals, significantly reduced tonic seizures and completely blocked mortality caused by PTZ but did not significantly modify the latency to the first convulsion. The results suggest that serotonin plays an important role in the protective effect of 0.5 mg/kg clonidine against PTZ-induced seizures. Possible reasons for the different effects of clonidine on different experimental seizures are discussed.     

Effects of bicuculline-induced seizures on benzodiazepine and adenosine receptors in developing rat brain

The effects of seizures induced by an acute administration of bicuculline have been investigated on the central benzodiazepine and adenosine receptors in developing rats and in adults. Generalized seizures rapidly increased the total number of both benzodiazepine binding sites and adenosine A1 receptors, without changes in receptor affinity (KD). It was concluded that such a phenomenon may facilitate the anticonvulsant action of benzodiazepine and adenosine via receptor binding and that it could be an adaptative process to protect subjects against recurrent seizures, especially in newborns.     

Osmoprotection by pipecolic acid in Sinorhizobium meliloti: specific effects of D and L isomers

DL-Pipecolic acid (DL-PIP) promotes growth restoration of Sinorhizobium meliloti cells facing inhibitory hyperosmolarity. Surprisingly, D and L isomers of this imino acid supplied separately were not effective. The uptake of L-PIP was significantly favored in the presence of the D isomer and by a hyperosmotic stress. Chromatographic analysis of the intracellular solutes showed that stressed cells did not accumulate radiolabeled L-PIP. Rather, it participates in the synthesis of the main endogenous osmolytes (glutamate and the dipeptide N-acetylglutaminylglutamine amide) during the lag phase, thus providing a means for the stressed cells to recover the osmotic balance. (13)C nuclear magnetic resonance analysis was used to determine the fate of D-PIP taken into the cells. In the absence of L-PIP, the imported D isomer was readily degraded. Supplied together with its L isomer, D-PIP was accumulated temporarily and thus might contribute together with the endogenous osmolytes to enhance the internal osmotic strength. Furthermore, it started to disappear from the cytosol when the L isomer was no longer available in the culture medium (during the late exponential phase of growth). Together, these results show an uncommon mechanism of protection of osmotically stressed cells of S. meliloti. It was proved, for the first time, that the presence of the two isomers of the same molecule is necessary for it to manifest an osmoprotective activity. Indeed, D-PIP seems to play a major role in cellular osmoadaptation through both its own accumulation and improvement of the utilization of the L isomer as an immediate precursor of endogenous osmolytes.     

Quantitative determination and regional distribution of pipecolic acid in rodent brain

A rapid and sensitive method for the quantitative determination of pipecolic acid (PA), one of the three cyclic secondary imino acids present in mammalian brain is described. The quantification and identification of PA are accomplished in rat and mouse brain using high performance liquid chromatography with electrochemical detection (LCEC) and nipecotic acid (NPA) as an internal standard. The cyclic imino acids are derivatized with 2,4-dinitrofluorobenzene (DNFB) to dinitrophenyl derivatives. The remaining time for LCEC analysis is less than 30 min and the limit of sensitivity is in the lower picomole range. The levels of PA found in rat and mouse brain are comparable to those reported using gas chromatography/mass spectrometry. The regional distribution of PA shows higher concentrations of PA in hypothalamus, pons-medulla oblongata and cerebellum. The present results demonstrate that LCEC is sensitive enough to determine endogenous levels of PA in mg amounts of rodent brain tissue. Due to its simplicity and rapidity, the technique represents an alternative to existing methods. This method can also be used for determination of PA in CSF, blood or urine of hyperipecolic patients.