Superimposing Status Epilepticus on Neuron Subset-Specific PTEN Haploinsufficient and Wild Type Mice Results in Long-term Changes in Behavior

Abstract

We evaluated the effects of superimposing seizures on a genetic mutation with known involvement in both Autism Spectrum Disorder and in epilepsy. Neuron-subset specific (NS)-Pten heterozygous (HT) and wildtype (WT) adult mice received either intraperitoneal injections of kainic acid (20 mg/kg) to induce status epilepticus or the vehicle (saline). Animals then received a battery of behavioral tasks in order to evaluate activity levels, anxiety, repetitive-stereotyped behavior, social behavior, learning and memory. In the open field task, we found that HT mice after seizures showed a significant increase in total activity and total distance in the surround region of the open field. In the elevated plus maze task, we found that HT mice after seizures displayed increased total distance and velocity as compared to HT mice that did not undergo seizures and WT controls. In the social chamber test, we found the HT mice after seizures displayed an impairment in social behavior. These findings demonstrate that superimposing seizures on a genetic mutation can result in long-term alterations in activity and social behavior in mice.

Figure 1. Status epilepticus in NS-Pten HT mice resulted in hyperactivity in the open field.

The mice were placed in an open field test for thirty minutes and several measurements of locomotor behavior were taken. (A) Total distance, (B) time performing stereotypy behavior, (C) frequency of stereotypy events, (D) number of rearing events, (E) number of clockwise rotations, and (F) number of counterclockwise rotations. The data were then reanalyzed by center and surround regions of the open field apparatus. (G) Total distance in the center and surround region of the open field test. Data are shown as mean ± standard error of the mean. **P > 0.01. WT-control n = 17, HT-control n = 12, WT-Seizure n = 14, HT-Seizure n = 14.

Figure 2. Status epilepticus in the NS-Pten HT mice resulted in hyperactivity without changes in anxiety in the elevated plus maze test.

All groups of mice were placed in a 10 minute elevated plus maze test. We first examined (A) overall changes in total distance in the maze and (B) velocity. (C,D) Duration and frequency of visits in the open, center, and closed arms in the maze. Separate video scoring was conducted for the (E,F) frequency and duration of head dips and (G,H) frequency and duration of rearing behavior in the closed arms. Data are shown as mean ± standard error of the mean. *P > 0.05. WT-control n = 18, HT-control n = 12, WT-Seizure n = 15, HT-Seizure n = 14.

Figure 3. No difference in repetitive behavior was found between Pten wildtype and heterozygous mice after status epilepticus.

There were no changes in the number of marbles buried in a 30 minute test. The number of marbles buried were examined at different levels: 50%, 75%, 100%, and completely buried. Data are shown as mean ± standard error of the mean. WT-control n = 18, HT-control n = 12, WT-Seizure n = 15, HT-Seizure n = 14.

Figure 4. Status epilepticus in the NS-Pten HT mice resulted in social behavior deficits.

(A) Time in the left, center, and right chamber in phase A. (B) Duration of interaction at the cups within the chamber in phase A. (C) Time in the chamber with the mouse, center, and chamber that housed the novel object. (D) Time spent with the cup that housed the mouse and object in the three chamber social behavior test. (E) Mean time interacting with the mouse compared to the novel object across groups. The values represent the mean ± SEM. ^#P = 0.05; *P < 0.05; **P < 0.01; ***P < 0.001. WT-control n = 18, HT-control n = 12, WT-Seizure n = 15, HT-Seizure n = 14.

Figure 5. Status epilepticus in the HT group resulted in enhanced learning in the trace fear conditioning test.

Mice were examined for differences in learning and memory through a trace fear conditioning test. (A) The HT mice with status epilepticus had more freezing across all trials. (B) No differences were observed in contextual conditioning a day later. Data are shown as mean ± standard error of the mean. *P < 0.05. WT-control n = 18, HT-control n = 12, WT-Seizure n = 15, HT-Seizure n = 14.

Figure 6. There were no spatial learning deficits after status epilepticus in the Morris water maze.

The mice were trained over eight blocks to find the hidden platform. There were no impairments to find the hidden platform (A). After 8 blocks of learning were completed, the animals were tested for spatial memory retention by the probe trial. There were no differences in the amount of time in each quadrant (B) or in the number of crossings in each zone (C). There were no differences in visible platform testing (D). Data are shown as mean ± standard error of the mean. WT-control n = 18, HT-control n = 12, WT-Seizure n = 15, HT-Seizure n = 14.