**6.2 Autosomal Dominant Nocturnal Frontal Lobe Epilepsy**

Autosomal Dominant Nocturnal Frontal Lobe Epilepsy (ADNFLE) is a focal epilepsy characterized by clusters of brief nocturnal motor seizures with hyperkinetic or tonic manifestations. Seizures typically occur very soon after falling asleep or during the early morning hours, initiated during stage 2 non-rapid-eye-movement (NREM) sleep. Onset usually occurs around age 10 and the seizures often persist through adult life. However, in most patients the seizures tend to peter out in adulthood.

In 1995, a missense mutation (S248F) in the nAChR α4 subunit gene (CHRNA4) was found to underlie ADNFLE (Steinlein et al., 1995). Additional disease causing mutations in this gene (Hirose et al., 1999; Steinlein et al., 1997), and mutations in the β2 subunit gene (CHRNB2) (Phillips et al., 2001) have later been identified. Because α4 and β2 subunits combine, it is not strange that mutations in either subunit produce comparable epileptic symptoms. Almost every known mutation is found in the TM2 region of the two subunits. A common finding among the mutations is that the sensitivity to ACh is increased (Bertrand et al., 2002).

### **6.3 How mutations in nAChRs can cause ADNFLE**

Stage 2 NREM sleep is characterized by the appearance of sleep spindles and slow waves, transient physiological rhythmic oscillations that are produced by synchronized synaptic potentials in cortical neurons. It appears that the seizure often arise from a sleep spindle that transforms into epileptic discharges (Picard et al., 2007). Thalamocortical circuits are thought to play a role in the generation of these sleep spindles.

Thalamic relay neurons are reciprocally connected to cortical neurons by excitatory synapses. Both thalamic and cortical neurons also excite GABAergic interneurons of the nucleus reticularis, which in turn inhibit thalamic relay neurons thus forming a feedbackloop (Kandel et al., 2000). The thalamic relay neurons have two different modes of signalling activity: a transmission mode during wakefulness and rapid eye movement (REM) sleep and a burst mode during NREM sleep. In the transmission mode, the resting membrane potential of the thalamic relay neurons is near the firing threshold and incoming excitatory synaptic potentials drive the neuron to fire in a pattern that reflects the sensory stimuli. During the burst mode the thalamic relay neurons are hyperpolarized and respond to brief depolarization with a burst of action potentials, which indicates that the thalamus is unable to relay sensory information to the cortex (Kandel et al., 2000). The thalamic relay neurons can be in different modes because they possess special Ca2+ channels named T-type channels. These channels are inactivated at the resting membrane potential but become available for activation when the cell is hyperpolarized, and can then be transiently opened by depolarization (Contreras, 2006). The interneurons of the nucleus reticularis that form synapses at the relay neurons hyperpolarize the relay neurons upon activation of GABAA receptors, thus removing the inactivation of the T-type Ca2+ channels. Incoming excitatory synaptic potentials can then trigger transient opening of the T-type Ca2+ channels and the

Epileptic Channelopathies and Dysfunctional

**7. Ion channel modulators as antiepileptic drugs** 

increase the sensitivity to the drug (Ortells and Barrantes, 2002).

sodium channel openers generally are known as epileptogenic substances.

An understanding of the etiology of the epilepsy syndromes and the causal factors in individual patients are also critical for selection of the right medication. Sodium channel blockers, which normally would exhibit anticonvulsant properties, would clearly not be the right choice to treat patients with SMEI, where a lack of sodium current seems to be the underlying cause. Similarly, as too much inhibition seems to play a role in the pathogenesis

Excitability - From Gene Mutations to Novel Treatments 203

Many of the antiepileptic drugs on the market today exert strong effects on ionic currents. Sodium channel blockers were used in the treatment of epilepsy as early as in 1940, and since then several other sodium channel blockers have been developed. Carbamazepine, first introduced in 1968, stabilizes the inactive conformation of sodium channels and is widely used in the treatment of partial and generalized tonic-clonic seizures. Lamotrigine blocks sodium channels in a voltage- and use-dependent manner and is efficient for partial, absence, myoclonic and tonic-clonic seizures (Kwan et al., 2001). Drugs that potentiate GABA receptor function, such as benzodiazepines, have also been used as anticonvulsants for several years. These act by binding to the interface between the α and γ subunits of the GABAA receptor, resulting in allosteric activation of the receptor (Kwan et al., 2001). Several established and novel antiepileptic drugs have been reported to act on various K+ currents, but none of them exert their main effect on potassium channels (Meldrum and Rogawski, 2007). Carbamazepine also exerts an effect on nAChRs by blocking the open conformation of the channel and is very effective in treatment of patients with ADNFLE. This effect might be related to the fact that several of the disease causing mutations in CHRNA4 and CHRNA2

The knowledge of how mutations in ion channels cause neuronal hyperexcitability and epilepsy has led to new therapeutic strategies for prevention of seizures. The mutated proteins serve as mechanistic proof of concept that pharmacological antagonization of the epilepsy causing mechanisms could have desirable effects on neuronal excitability. These mechanisms are not necessarily targeted for treatment of the respective epilepsy syndrome, but as they are showed to cause hyperexcitability in the affected patients, antagonizing drugs may reduce excitability in patients with other types of epilepsy. An excellent example of this is Retigabine (other names are Trobalt, Potiga, or Ezogabine), which was approved by both the European Medicines Agency and the U.S. Food and Drug Administration in 2011 as adjunctive treatment for partial onset seizures. It was originally synthesized as a GABA modulator, but it was later shown that its main molecular target is the neuronal Kv7 channels (Main et al., 2000; Rundfeldt and Netzer, 2000; Wickenden et al., 2000). Retigabine induces a large hyperpolarizing shift in the voltage-dependence of activation of Kv7 channels, accelerates the activation kinetics and slows deactivation kinetics. Enhancement of M-current would clearly be effective in prevention of seizures in BNFC patients, but mutations in the Kv7 channels might render them insensitive to such drugs, which hampers the use of Kv7 channel openers in the treatment of BNFC. The hypothesis of reduced NaV1.1 current in the pathogenesis of GEFS+ and SMEI, together with its primary expression in inhibitory interneurons, indicates that this sodium channel subtype plays a role in reducing neuronal excitability (Ogiwara et al., 2007; Yu et al., 2006). This opens the intriguing possibility that selective NaV1.1 openers can function as anticonvulsants, even though

resulting influx of Ca2+ brings the neuron's membrane potential above threshold. The cell now fires a burst of action potentials that produces the synchronized postsynaptic potentials in cortical neurons that cause the spindle waves seen on the electroencephalogram. When sufficient Ca2+ has entered the cell a Ca2+-activated K+ current is triggered that hyperpolarizes the relay neurons and terminate the spindle (Contreras, 2006). Because of the feedback loops from the relay neurons and cortical neurons that innervate the interneurons of the nucleus reticularis, these are again activated, which allows for another round of burst firing.

There is a high expression of the α4β2 nAChR subtype in the thalamus and the subtype can be found diffusely distributed onto pyramidal cells and GABAergic interneurons in the cortex. The thalamic relay neurons and the nucleus reticularis receive input from two groups of cholinergic neurons in the upper brain stem: the pedunculopontine and laterodorsal tegmental nuclei (LDT). These are critical for keeping the thalamic relay neurons in transmission mode during wakefulness and REM sleep. The LDT releases Ach during NREM sleep at the time of an arousal that interrupts the sleep spindle oscillations through depolarization of thalamic relay neurons (Lee and McCormick, 1997). The cortex receives cholinergic input from neurons in the nucleus basalis of Meynert that enhance cortical response to incoming sensory stimuli (Kandel et al., 2000).

After studying transgenic ADNFLE mice with heterozygous expression of an ADNFLE mutation (Chrna4S252F or Chrna4+264), Boulter and colleagues suggested that an Achdependent sudden increase in the response of GABAergic cortical interneurons contributes to the epileptogenesis in ADNFLE patients (Klaassen et al., 2006). They showed that asynchronously firing cortical pyramidal cells got synchronized when relived from a large GABAergic inhibition triggered by cholinergic over-activation of mutant nAChRs in the cortical interneurons.

Dani and Bertrand suggested that this, together with an increased positive response of cortical pyramidal cells through cholinergic stimulation of their hypersensitive α4β2 nAChRs, contributes to induction of the seizures (Dani and Bertrand, 2007). As the cortical pyramidal cells project back to the thalamus, excitatory stimuli will be boosted on to the interneurons of the nucleus reticularis and the relay neurons and induce synchronous activity and seizures (Dani and Bertrand, 2007).

In addition, PET studies using a high affinity α4β2 agonist have showed a clear difference in the pattern of the nAChR density in the brains of ADNFLE patients compared to control subjects (Picard et al., 2006). The studies showed that patients had increased α4β2 density in the epithalamus, the interpeduncular nucleus (IPN) of the ventral mesencephalon and in the cerebellum, while the density in the right dorsolateral prefrontal region was decreased. As the IPN projects to the LDT, the authors proposed that prolonged depolarizations in IPN neurons, because of both increased density of α4β2 and hypersensitive receptors, could result in over-activation of the LDT and consequently the thalamic relay neurons (Picard et al., 2006). At the time of arousals, ACh acting on sensitized thalamic relay neurons could prevent the normal arousal-induced interruption of the sleep spindle oscillations and transform them into pathological Thalamocortical oscillations, triggering epileptic seizures (Picard et al., 2006).

Neither of these mechanisms excludes the others. In fact, they might work together yielding even stronger possibilities for induction of seizures.

resulting influx of Ca2+ brings the neuron's membrane potential above threshold. The cell now fires a burst of action potentials that produces the synchronized postsynaptic potentials in cortical neurons that cause the spindle waves seen on the electroencephalogram. When sufficient Ca2+ has entered the cell a Ca2+-activated K+ current is triggered that hyperpolarizes the relay neurons and terminate the spindle (Contreras, 2006). Because of the feedback loops from the relay neurons and cortical neurons that innervate the interneurons of the nucleus reticularis, these are again activated, which allows for another round of burst

There is a high expression of the α4β2 nAChR subtype in the thalamus and the subtype can be found diffusely distributed onto pyramidal cells and GABAergic interneurons in the cortex. The thalamic relay neurons and the nucleus reticularis receive input from two groups of cholinergic neurons in the upper brain stem: the pedunculopontine and laterodorsal tegmental nuclei (LDT). These are critical for keeping the thalamic relay neurons in transmission mode during wakefulness and REM sleep. The LDT releases Ach during NREM sleep at the time of an arousal that interrupts the sleep spindle oscillations through depolarization of thalamic relay neurons (Lee and McCormick, 1997). The cortex receives cholinergic input from neurons in the nucleus basalis of Meynert that enhance

After studying transgenic ADNFLE mice with heterozygous expression of an ADNFLE mutation (Chrna4S252F or Chrna4+264), Boulter and colleagues suggested that an Achdependent sudden increase in the response of GABAergic cortical interneurons contributes to the epileptogenesis in ADNFLE patients (Klaassen et al., 2006). They showed that asynchronously firing cortical pyramidal cells got synchronized when relived from a large GABAergic inhibition triggered by cholinergic over-activation of mutant nAChRs in the

Dani and Bertrand suggested that this, together with an increased positive response of cortical pyramidal cells through cholinergic stimulation of their hypersensitive α4β2 nAChRs, contributes to induction of the seizures (Dani and Bertrand, 2007). As the cortical pyramidal cells project back to the thalamus, excitatory stimuli will be boosted on to the interneurons of the nucleus reticularis and the relay neurons and induce synchronous

In addition, PET studies using a high affinity α4β2 agonist have showed a clear difference in the pattern of the nAChR density in the brains of ADNFLE patients compared to control subjects (Picard et al., 2006). The studies showed that patients had increased α4β2 density in the epithalamus, the interpeduncular nucleus (IPN) of the ventral mesencephalon and in the cerebellum, while the density in the right dorsolateral prefrontal region was decreased. As the IPN projects to the LDT, the authors proposed that prolonged depolarizations in IPN neurons, because of both increased density of α4β2 and hypersensitive receptors, could result in over-activation of the LDT and consequently the thalamic relay neurons (Picard et al., 2006). At the time of arousals, ACh acting on sensitized thalamic relay neurons could prevent the normal arousal-induced interruption of the sleep spindle oscillations and transform them into pathological Thalamocortical oscillations, triggering epileptic seizures

Neither of these mechanisms excludes the others. In fact, they might work together yielding

cortical response to incoming sensory stimuli (Kandel et al., 2000).

firing.

cortical interneurons.

(Picard et al., 2006).

activity and seizures (Dani and Bertrand, 2007).

even stronger possibilities for induction of seizures.
