Neuronal inhibition and seizure suppression by acetoacetate and its analog, 2-phenylbutyrate.
OBJECTIVE
The ketogenic diet is clinically used to treat drug-resistant epilepsy. The diet treatment markedly increases ketone bodies (acetoacetate and β-hydroxybutyrate), which work as energy metabolites in the brain. Here, we investigated effects of acetoacetate on voltage-dependent Ca2+ channels (VDCCs) in pyramidal cells of the hippocampus. We further explored an acetoacetate analog that inhibited VDCCs in pyramidal cells, reduced excitatory postsynaptic currents (EPSCs), and suppressed seizures in vivo.


METHODS
The effects of acetoacetate and its analogs on VDCCs and EPSCs were evaluated using patch-clamp recordings from CA1 pyramidal cells of mouse hippocampal slices. The in vivo effects of these reagents were also evaluated using a chronic seizure model induced by intrahippocampal injection of kainate.


RESULTS
Acetoacetate inhibited VDCCs in pyramidal cells of hippocampal slices, and reduced EPSCs in slices exhibiting epileptiform activity. More potent EPSC inhibitors were then explored by modifying the chemical structure of acetoacetate, and 2-phenylbutyrate was identified as an acetoacetate analog that inhibited VDCCs and EPSCs more potently. Although acetoacetate is known to inhibit vesicular glutamate transporters (VGLUTs), 2-phenylbutyrate did not inhibit VGLUTs, showing that 2-phenylbutyrate is an acetoacetate analog that preferably inhibits VDCCs. In addition, 2-phenylbutyrate markedly reduced EPSCs in slices exhibiting epileptiform activity, and suppressed hippocampal seizures in vivo in a mouse model of epilepsy. The in vivo antiseizure effects of 2-phenylbutyrate were more potent than those of acetoacetate. Finally, intraperitoneal 2-phenylbutyrate was delivered to the brain, and its brain concentration reached the level enough to reduce EPSCs.


SIGNIFICANCE
These results demonstrate that 2-phenylbutyrate is an acetoacetate analog that inhibits VDCCs and EPSCs in pyramidal cells, suppresses hippocampal seizures in vivo, and has brain penetration ability. Thus 2-phenylbutyrate provides a useful chemical structure as a lead compound to develop new antiseizure drugs originating from ketone bodies.
['Kadowaki|Atsushi|A|', 'Sada|Nagisa|N|', 'Juge|Narinobu|N|', 'Wakasa|Ayaka|A|', 'Moriyama|Yoshinori|Y|', 'Inoue|Tsuyoshi|T|']
D000090:Acetoacetates D000818:Animals D015220:Calcium Channels D055423:Diet, Ketogenic D004195:Disease Models, Animal D004569:Electroencephalography D005260:Female D006624:Hippocampus D066298:In Vitro Techniques D007267:Injections D007608:Kainic Acid D008297:Male D051379:Mice D008813:Mice, Inbred ICR D009433:Neural Inhibition D009924:Organ Culture Techniques D010654:Phenylbutyrates D017966:Pyramidal Cells
Ketogenic diet Voltage-dependent Ca2+ channel Hippocampus Antiseizure effect Ketone body analog