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Keywords:

  • Bumetanide;
  • Temporal lobe epilepsy;
  • NKCC1;
  • GABA

Summary

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

Alterations in the balance of K-Na-2Cl cotransporter (NKCC1) and Na-Cl cotransporter (KCC2) activity may cause depolarizing effect of γ-aminobutyric Acid (GABA), and contribute to epileptogenesis in human temporal lobe epilepsy. NKCC1 facilitates accumulation of chloride inside neurons and favors depolarizing responses to GABA. In the current pilot study we provide the first documented look at efficacy of bumetanide, a specific NKCC1 antagonist, on reduction of seizure frequency in adult patients with temporal lobe epilepsy. According to our results, seizure frequency was reduced considerably in these patients. Furthermore, epileptiform discharges decreased in two of our patients. If the efficacy of bumetanide is proven in large scale studies, it can be used as a supplemental therapy in temporal lobe epilepsy.

It has been shown that changes in γ-aminobutyric acid (GABA)ergic signaling play an important role in temporal lobe epileptogenesis (Cohen et al., 2003). GABA, the main inhibitory transmitter in the adult central nervous system (CNS), at early developmental stages depolarizes target cells through an outwardly directed flux of chloride. In mature neurons, because of the low level of intracellular chloride ([Cl]i), GABA triggers membrane hyperpolarization due to passive influx of chloride down its electrochemical gradient. In contrast, immature neurons have an elevated intracellular [Cl]i, so that GABA triggers chloride efflux and membrane depolarization (Ben-Ari, 2002). Two cation-chloride cotransporters may be especially important in controlling neuronal intracellular [Cl] the Na-K-2Cl cotransporter (NKCC1) loads neurons with Cl and favors depolarizing responses to GABA, whereas the K-Cl cotransporter (KCC2) normally extrudes Cl ions, thereby promoting hyperpolarizing responses (Ben-Ari et al., 2007). In patients with epilepsy, changes in expression of NKCC1 and KCC2 may cause a depolarizing effect of GABA and contribute to epileptogenesis in the hippocampal formation (Ben-Ari & Holmes, 2005).

According to previous studies, patients with temporal lobe epilepsy (TLE) showed depolarizing GABAergic neurons in subicular pyramidal cells, and bumetanide suppressed this activity (Huberfeld et al., 2007). Muñoz et al. (2007), showed upregulation of the NKCC1 mRNA in the human epileptic hippocampus. Bumetanide is a specific NKCC1 antagonist that reduces [Cl]i, and switches GABA from excitation to inhibition (Delpire & Mount, 2002). This drug has been used in adults as a diuretic since 1975, with known pharmacokinetics and side effects (Sullivan et al., 1996). Suppression of seizure activity by bumetanide in a neonate was studied by Kahle et al. (2009).

In the current study, we provide the first documented data of bumetanide efficacy on reduction of seizure frequency in adult patients with temporal lobe epilepsy (TLE).

Methods

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

We evaluated the effect of bumetanide in patients with drug-resistant TLE who were candidates for surgery. This study has been approved by the ethical committee of Tehran University of medical sciences, and all patients gave a written informed consent prior to start this study.

Patients

Patients were selected according to the following criteria.

  • Inclusion criteria:

    • Having more than three seizure attacks followed by altered consciousness in the last 3 months before the first visit.

    • History of automotor or dialeptic seizures/epigastric or psychiatric aura.

    • Electroencephalography (EEG) changes:

      • TIRDA (temporal intermittent rhythmic delta activity), or

      • Interictal spikes over F7, F8, F9, F10, FT7, FT8, FT9, FT10, or

      • Ictal rhythmic activity over F7, F8, F9, F10, FT7, FT8, FT9, FT10 (EEG changes should be started before clinical symptoms or 5 s after clinical symptoms at the latest).

    • Magnetic resonance imaging (MRI) changes: Increased hippocampal T2 signal or hippocampal or fornix atrophy.

  • Exclusion criteria:

    • History of trauma, encephalopathy, hypoxia, diffuse, or extratemporal focal EEG spikes or other cerebral lesions (Table S1).

Study design

According to inclusion criteria, three patients were selected. All patients were followed before, during, and after treatment for 4, 4, and 3 months respectively. Bumetanide was administered 2 mg daily. Preexisting antiepileptic treatment of patients remained unchanged. We monitored for side effects of bumetanide by doing blood serum analysis before and every month during treatment. All patients were visited weekly to monitor their symptoms and quality of life. Seizure and aura days in this study referred to the number of days in which seizure and aura happened regardless of the total number of seizures.

Long-term video-EEG monitoring (LTM)

Long-term video-EEG monitoring (LTM) (24 h monitoring) was done using Stellate LTM System (Stellate, Montreal, QC, Canada) (Fig. S1) before and after treatment.

Results

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

Effects of bumetanide on seizure frequency

The first patient was a 37-year-old man with symptomatic epilepsy and seizure onset from the left temporal lobe (Table 1). Seizures were characterized by epigastric aura followed by an impairment of consciousness and motor automatisms. The patient showed a seizure-day frequency reduction of 68% (47 seizure-days/4 months before treatment compared to 15 seizures-days/4 months after treatment initiation).

Table 1.   Number of days with seizure/aura before and after treatment initiation
Patient4 Months before treatment initiation4 Months after treatment initiation
No. days with only auraNo. seizure days ± aurasNo. days with only auraNo. seizure days ± auras
  1. aAttacks exclusively happened after stressful situation and fatigue.

1047015
201372a
30401a

The second patient was a 31-year-old man with symptomatic epilepsy and seizure onset from the left temporal lobe. His seizure attacks were in the form of auditory illusion and sometimes déjà vu followed by staring, left cheek jerks, asymmetrical smile and occasional salivation, right hand automatism, and postictal confusion without any secondary generalized seizures. The patient showed a seizure-day frequency reduction of 84% (13 seizure-days/4 months before treatment compared to 2 seizure-days/4 months after treatment initiation). During treatment period most of his attacks were just auras, and he had only two seizures accompanied by consciousness alteration; both happened after very stressful situations. In the pretreatment period his wife could recognize his seizures, but during treatment the attacks were too weak to be noticed by his wife. His self-confidence improved, and he was able to take more career-related travel.

The third patient was a 32-year-old man with symptomatic epilepsy and seizure onset from the left anterior temporal lobe. His seizures were characterized by impairment of consciousness, oral and right hand automatisms, and a short postictal phase. The patient showed a seizure-day frequency reduction of 75% (four seizure days/4 months before treatment compared to one seizure day/4 months after treatment initiation). The posttreatment seizure happened after a very stressful situation.

Patient satisfaction

We evaluated subjective patient-judged rating of satisfaction with bumetanide treatment. Patients 2 and 3 were very satisfied with the treatment and interested in pursuing it (Table S2).

Long-term video-EEG monitoring

The number of interictal discharges was evaluated (Table 2). Second and third patients showed a reduction of 100% (three interictal discharges in pretreatment LTM compared to no interictal discharge in posttreatment LTM) and 67.5% (74 interictal spikes in pretreatment LTM compared to 24 interictal spike in posttreatment LTM), respectively. Interictal discharge frequency increased for the first patient after treatment (25 interictal discharge in pretreatment LTM compared to 38 interictal spike in posttreatment LTM).

Table 2.   Long-term video-EEG monitoring results before and after treatment initiation
PatientLTM before treatment initiationLTM after treatment initiation
No. interictal dischargesNonepileptiform dischargesNo. interictal dischargesNonepileptiform discharges
125Rare38Rare
23Rare0Rare
374Frequent24Moderate

We scored nonepileptiform discharge occurrence as follows: Nil, 0; Rare, 1–10; Moderate, 10–100; Frequent, >100 (Table 2; Figs S2–S4).

Posttreatment follow-up

Within the 3-month posttreatment follow-up, Patients 2 and 3 remained completely seizure free, but the first patient had about 11 seizure days in this period.

Discussion

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

Epilepsy with debilitating effects on health and lifestyle, has a high psychological impact (van Andel et al., 2011), and any beneficial treatment can positively improve quality of life in patients.

In a study by Dzhala et al. (2008) on newborn rats, phenobarbital in combination with bumetanide had an anticonvulsant effect on seizure frequencies. A study by Muñoz et al. (2007) on human epileptic tissues revealed that NKCC1 is up-regulated, whereas KCC2 is down-regulated in some subicular cells in patients with epilepsy. In another study done by Huberfeld et al. (2007), application of bumetanide on human TLE slices suppressed interictal-like activity and restored hyperpolarizing GABAergic responses.

In our study we had three adult patients with drug-resistant TLE who received bumetanide. We noticed considerable reduction in their number of seizure days. It is noteworthy to mention that during the treatment period, two of our patients (Patients 2 and 3) had seizures exclusively following stressful situations, and they were completely seizure-free after treatment follow-up. Positive changes such as being able to take more career-related travel, increase working hours, and improved self-confidence were reported by these patients. Furthermore, according to the LTM results, supplementation of bumetanide caused a significant reduction in epileptiform and nonepileptiform discharges in Patient 3. Interictal discharges in the second patient decreased as well. These two patients were very satisfied with the treatment and interested in pursuing it. As Kahle et al. (2009) mentioned in their study in human infants, it is not clear whether this seizure frequency reduction is due to effects of bumetanide on NKCC1 action, or only a coincidence. On the other hand, according to previous studies, loop diuretics by modulating the size of extracellular space can block epileptiform activity in adult brain (Hochman, 2012). In contrast, there are several lines of evidence indicating that bumetanide exerts antiepileptic effects in neonatal seizure via synaptic mechanisms related to excitatory GABAergic currents (Dzhala et al., 2008; Kahle et al., 2009). In addition, (Huberfeld et al., 2007) have shown that bumetanide through GABAergic signaling reduced interictal activity without changing extracellular volume. When they induced interictal activity in the subiculum of slices from epileptic patients by increasing K+ in the presence of picrotoxin, this activity, which is independent of GABAA receptors, was not suppressed by bumetanide application.

The results obtained in this study revealed that bumetanide can be a suitable substitute for surgical treatment of patients with nonresponsive TLE. More extensive studies will be needed to elucidate the exact role of bumetanide in seizure control. We are establishing a multicenter study in order to show the exact effects of this drug.

If the efficacy of bumetanide be proven in large-scale studies, it will be used as a supplement therapy in temporal lobe epilepsies, or even as a substitute for surgery.

Acknowledgments

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

The authors of this manuscript would like to acknowledge their thanks to the grant no. 508 offered by cellular and molecular research centre of Tehran University of medical sciences. In addition, we want to thank Iran National Science Foundation (INSF) for their support.

Disclosure

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

None of the authors has any conflict of interest to disclose. We confirm that we have read the Journal’s position on issues involved in ethical publication and affirm that this report is consistent with those guidelines.

References

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

Supporting Information

  1. Top of page
  2. Summary
  3. Methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. Disclosure
  8. References
  9. Supporting Information

Figure S1. The location of electrodes is illustrated (31 active electrodes, two pre-cordial electrodes, one earth electrode and one reference electrode).

Figure S2. Illustrates samples from long-term video EEG monitoring of Patient 1.

Figure S3. Illustrates samples from long-term video EEG monitoring of Patient 2.

Figure S4. Illustrates samples from long-term video EEG monitoring of Patient 3.

Table S1. Summary of patients included in this study.

Table S2. Patient satisfaction.

FilenameFormatSizeDescription
epi3654_sm_figureS1.tif1742KSupporting info item
epi3654_sm_figureS2.tif6349KSupporting info item
epi3654_sm_figureS3.tif6884KSupporting info item
epi3654_sm_figureS4.tif6686KSupporting info item
epi3654_sm_TableS1.docx14KSupporting info item
epi3654_sm_TableS2.docx13KSupporting info item

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