Case Report of Novel Genetic Variant in KCNT1 Channel and Pharmacological Treatment With Quinidine. Precision Medicine in Refractory Epilepsy.

Case introduction: In this work we present a female infant patient with epilepsy of infancy with migrating focal seizures (EIMFS). Although many pharmacological schemes were attempted, she developed an encephalopathy with poor response to antiepileptic drugs and progressive cerebral dysfunction. Aim: To present the pharmacological response and therapeutic drug monitoring of a paediatric patient with a severe encephalopathy carrying a genetic variant in KCNT1 gene, whose identification led to include quinidine (QND) in the treatment regimen as an antiepileptic drug. Case report: Patient showed slow rhythmic activity (theta range) over left occipital areas with temporal propagation and oculo-clonic focal seizures and without tonic spasms three months after birth. At the age of 18 months showed severe impairments of motor and intellectual function with poor eye contact. When the patient was 4 years old, a genetic variant in the exon 24 of the KCNT1 gene was found. This led to the diagnosis of EIMFS. Due to antiepileptic treatment failed to control seizures, QND a KCNT1 blocker, was introduced as a therapeutic alternative besides topiramate (200 mg/day) and nitrazepam (2 mg/day). Therapeutic drug monitoring (TDM) of QND plasma levels needed to be implemented to establish individual therapeutic range and avoid toxicity. TDM for dose adjustment was performed to establish the individual therapeutic range of the patient. Seizures were under control with QND levels above 1.5 mcg/ml (65-70 mg/kg q. i.d). In addition, QND levels higher than 4.0 mcg/ml, were related to higher risk of suffering arrhythmia due to prolongation of QT segment. Despite initial intention to withdrawal topiramate completely, QND was no longer effective by itself and failed to maintain seizures control. Due to this necessary interaction between quinidine and topiramate, topiramate was stablished in a maintenance dose of 40 mg/day. Conclusion: The implementation of Precision Medicine by using tools such as Next Generation Sequencing and TDM led to diagnose and select a targeted therapy for the treatment of a KCNT1-related epilepsy in a patient presented with EIMFS in early infancy and poor response to antiepileptic drugs. QND an old antiarrhythmic drug, due to its activity as KCNT1 channel blocker, associated to topiramate resulted in seizures control. Due to high variability observed in QND levels, TDM and pharmacokinetic characterization allowed to optimize drug regimen to maintain QND concentration between the individual therapeutic range and diminish toxicity.

Effects of clenbuterol treatment on the relaxant response in rat uterus.

Prolonged treatment with the beta(2)-adrenergic agonist clenbuterol (0.25 mg kg(-1) s.c. once daily for 10 days) produced a reduction of the relaxant response and cAMP production mediated by stimulation of beta-adrenoceptors in oestrogen-treated rat uterus. Substantial decreases in the relaxant effect of isoproterenol is observed in uterine rings precontracted with 50 mM KCl from clenbuterol-treated rats. The recovery of the relaxant response was also studied and significant differences were seen between acute and prolonged treatment with clenbuterol (P<0.05 vs control). In contrast the relaxant effect of forskolin and 3-isobutyl-1-methylxanthine was similar in untreated or treated rats. Sodium fluoride also showed a relaxant response which was not affected by the treatment with clenbuterol. The radioligand studies showed a reduction in the number of beta-adrenoreceptors after acute and prolonged treatment with clenbuterol in rat uterus. These results suggest that prolonged treatment with clenbuterol caused a desensitization of the relaxant uterine response through beta(2)-adrenoceptors and also showed differences in the recovery of the relaxant response depending on the duration of treatment.

Effects of optical isomers of clenbuterol on the relaxant response in rat uterus.

Previous works have shown that the administration of the racemic beta -adrenoceptor agent clenbuterol produces a desensitization of the relaxant response in rat uterus. The aim of this work was to study the effects of the optical isomers of clenbuterol on the relaxant response in rat uterus. The administration of (L)-clenbuterol (0.25 mg per kg per day) over 1 or 10 consecutive days, produced a reduction of the relaxant response to isoproterenol in uterine rings precontracted with 50 m m KCl from oestrogenic rats. The administration of (D)-clenbuterol (0.25 mg per kg per day) over 1 or 10 days did not affect the relaxant response of isoproterenol. (L)-clenbuterol also produced a concentration-dependent relaxant effect that was not observed with (D)-clenbuterol. These results show that the beta-adrenergic relaxant response and the desensitization of the relaxant effect to isoproterenol is due to the (L)-isomer and that the (D)-isomer is not involved in these effects.

[Effect of treatment with dexamethasone on cardiovascular responses of adrenergic agents]. / Efecto del tratamiento con dexametasona sobre las respuestas cardiovasculares de agentes adrenérgicos.

Cardiovascular responses to several agents should be modified by glucocorticoid administration in the rat. We investigate the response to adrenergic agonists such as phenylephrine, noradrenaline, clonidine and isoproterenol and ganglionic blocking agent such as hexamethonium in conscious rats treated during 7 days with dexamethasone. Wistar rats were treated with either Dex (150 micrograms daily x 7 days, p.o.) or water. Mean arterial pressure were calculated from the intraarterial recordings of blood pressure. No differences in basal mean arterial pressure were seen between dexamethasone and control groups of rats. Phenylephrine and noradrenaline showed a pressor effect in control rats that was reduced by dexamethasone treatment. Clonidine showed similar pressor effect in both groups of rats but ten minutes after drug administration, a light hypotension was seen in dexamethasone rats. Isoproterenol and hexamethonium showed a similar hypotensive effect on control and dexamethasone rats. In conclusion, dexamethasone treatment should reduce the pressor responses to phenylephrine and noradrenaline. Moreover, the alpha adrenergic agonist clonidine showed a hypotensive effect in dexamethasone treated rats, although the response of isoproterenol and hexamethonium remains unchanged.

Pharmacokinetic study of methyldopa in aorta-coarctated rats using a microdialysis technique.

The pharmacokinetics of methyldopa (12.5, 25 and 50 mg kg(-1), i.p.) was studied in anesthetized sham-operated (SO) and abdominal aorta-coarctated (ACo) rats using a microdialysis technique. A non-linear relationship between the area under the curve (AUC) and dose was observed in SO rats. However, in ACo rats the AUC showed a proportional increase with dose. Abdominal aortic coarctation produced significant differences in the estimates of clearance (Cl) and the elimination rate constant from the dialysate (K(ed)) after the administration of 50 mg kg(-1)of methyldopa (K(ed)SO, 0.31 +/- 0.09; ACo, 0.66 +/- 0.09(*)h(-1): Cl SO, 30.8 +/- 10.1; ACo, 78.6 +/- 13.3(*)mlkg(-1)min(-1);n= 6,(*)P< 0.05 vs SO). In conclusion, this study, by using a microdialysis technique, suggests that abdominal aortic coarctation seems to produce changes in the pharmacokinetics of methyldopa in rats.

Study of the evolution of blood and striatal levels of methyldopa: a microdialysis study in sinaortic denervated rats.

Pharmacokinetic of methyldopa (50 mg kg(-1)i.p.) was studied in anesthetized sham operated and sinoaortic denervated (SAD) rats by using the microdialysis technique. Vascular shunt probe was inserted into the carotid artery and concentric probe was placed in the striatum. Levels of methyldopa were measured by HPLC-EC. The number of animals in each group was six and normal distribution of the variables of the study was assumed. Peak concentrations in arterial blood of methyldopa were similar in both groups of rats but the elimination rate constant was 0.31+/-0.09 h(-1)for sham rats (n =6) and 1.28+/-0.31 h(-1)for SAD rats (n =6, P<0.05). Low levels of methyldopa and a more pronounced decrease were seen in striatum of sinoaortic denervated rats. In conclusion, by using a microdialysis technique, different kinetic profiles of methyldopa were observed in sham operated and sinoaortic denervated rats.

Beta-adrenoceptor desensitization by/clenbuterol in rat uterus.

1. The relaxant response and cAMP production mediated by stimulation of isoproterenol is reduced in uterine rings from clenbuterol treated rats (0.25 mg kg-1 s.c. 24 hr before experiments) precontracted with 50 mM KCl. 2. Forskolin, in contrast, showed similar relaxant responses in untreated or clenbuterol treated rats. 3. Isoproterenol produces a biphasic response that is composed of a rapid relaxation followed by a slower regaining of tension, which is considered as desensitization. 4. The kinetic study demonstrates marked changes in the desensitization process of beta-adrenoceptors after clenbuterol administration.

Pharmacokinetic and pharmacodynamic alterations of methyldopa in rats with aortic coarctation. A study using microdialysis.

A pharmacokinetic-pharmacodynamic study of methyldopa (MD) was made in anesthetized sham operated (SO) and aortic coarctated (ACo) rats by using a vascular shunt probe for arterial microdialysis and simultaneous blood pressure recording. Anesthetized Wistar rats were used 7 days after aortic coarctation or sham operation. A vascular shunt probe was inserted into the carotid artery and a concentric probe was placed into the striatum or posterior hypothalamus. MD and 3,4-dihydroxyphenylacetic acid (DOPAC) were determined in the dialysates by HPLC-EC. MD (50 mg kg(-1)i.p.) induced an increase of heart rate in SO (Delta HR: 108 +/- 22 bpm, n= 6) and in ACo rats (Delta HR: 55 +/- 10 bpm, n= 6, P< 0.05, one way ANOVA). Moreover, MD also reduced the mean arterial pressure (MAP) of SO rats (Delta MAP: -10 +/- 4 mmHg, n= 6) and ACo animals (Delta MAP: -51 +/- 9 mmHg, n= 6, P< 0.05, one way ANOVA). Analysis of the arterial blood dialysates showed a lower half-life of MD in ACo rats (t(1/2): 1.5 +/- 0.3 h, n= 6, P< 0.05, 't' test) than in SO rats (t(1/2): 3.7 +/- 1.0 h, n= 6). A low accumulation and a fast decay of striatal MD levels were seen in ACo rats. However, peak levels of drug were greater in the hypothalamic dialysates of ACo rats than in SO animals samples. On the other hand, MD also induced an increase of DOPAC levels in the hypothalamic dialysates of ACo rats. In conclusion, the aortic coarctation modifies the pharmacokinetic and cardiovascular effect of MD in the rat. The action of this drug on dopaminergic neurotransmission is also altered in the ACo animals.