Excitatory synaptic function is selectively enhanced in CA1 of hippocampal brain slices following <i>in vitro</i> SAHA treatment.

<p>(A) The median amplitude of mEPSCs was significantly increased in SAHA-treated slices (p<0.05, n = 14 vehicle, 14 SAHA), while there was no significant change to mEPSC frequency as measured by the median interval between events (p>0.05). Example mEPSC traces from vehicle and SAHA treated slices are shown inset (scale bar represents 10 pA and 250 ms). (B) SAHA treatment did not significantly alter the amplitude or frequency of mIPSCs (p>0.05, n = 14, 11). Example mIPSC traces from vehicle and SAHA treated slices are shown inset (scale bar represents 10 pA and 500 ms). All data points are plotted as mean ±SEM.</p

SAHA treated slices exhibit enhanced induction of LTP and impaired LTD.

<p>(A) An induction protocol that was subthreshold in vehicle treated slices readily evoked LTP in SAHA treated slices (p<0.05, n = 5,5). Example traces before and after LTP induction are shown in red for SAHA and black for vehicle treated slices (scale bars represent 20 pA and 20 ms). (B) An induction protocol that readily induced LTD in vehicle treated slices could not produce LTD in SAHA treated slices (p<0.05, n = 9 vehicle, 8 SAHA). Example traces before and after LTD induction are shown in red for SAHA and black for vehicle treated slices (scale bars represent 25 pA and 20 ms). Data are plotted as mean ±SEM.</p

Pharmacokinetic analysis of SAHA following i.p. injection.

<p>A) Bioanalysis of the time course of total (top) and unbound (bottom) plasma, CSF, and brain levels of SAHA following a single 50 mg/kg ip injection (n = 3 mice/time point). The dotted red lines represent the SAHA concentration imposed on the <i>in vitro</i> slice cultures for the electrophysiological studies. B) Total (top) and unbound (bottom) SAHA levels are shown following a 150 mg/kg ip injection (n = 3/time point). All data is shown as mean ± SD.</p

Fear memory deficits in Tg2576 mice are not rescued by SAHA treatment.

<p>A) Compared to non-transgenic littermates (n = 15), Tg2576 (n = 14) mice showed significantly less freezing than wt mice when returned to the context in which conditioning occurred (context, p<0.001), when placed in an altered context (altered, p<0.01), or in response to the cue used for conditioning (cue, p<0.05). B) Tg2576 mice were treated daily for 33 days prior to, as well as during fear conditioning with either vehicle (n = 14), 25 mg/kg SAHA (n = 13), or 50 mg/kg SAHA (n = 13). There was no effect of treatment on the percentage of time Tg2576 mice spent freezing in response to the context, altered context, or cue (p>0.05). C) There was no effect of treatment on the percentage of time spent freezing during conditioning (p>0.05). D) There was no effect of treatment on the distance traveled in the open field test (total distance = 38.0±7.7 m for vehicle, 39.6±6.1 m for 25 mg/kg SAHA, and 34.2±5.2 m for 50 mg/kg SAHA, p>0.05). All data are plotted as mean ±SEM.</p

SAHA IC₅₀ values for each HDAC isozyme are shown as measured using <i>in vitro</i> enzymatic assays.

<p>Values are in µM and confidence intervals are in parentheses.</p

Acute or chronic SAHA treatment does not produce significant drug class activity signatures as assessed by the SmartCube®.

<p>A. Groups of mice were treated acutely with a single injection of 50 mg/kg or 150 mg/kg SAHA or vehicle. In addition, a group was treated with valproate (225 mg/kg). Both does of SAHA were behaviorally inactive without a clear therapeutic signal. In contrast valproate was behaviorally active (p<0.001, discrimination index = 100%) with a strong anxiolytic signature and a mild psychostimulant signature. B. Groups of mice were treated daily for 14 days with SAHA or Valproate. While the lower dose of SAHA appeared behaviorally active (p<0.001, discrimination index = 88%), the activity was not consistent with any known therapeutic signal and the higher dose was not behaviorally active. In contrast valproate showed a strong behavioral activity (p<0.001, discrimination index = 98%) with a predominantly anxiolytic signature. C. The legend shows the 15 classes of behavioral activity that were assessed.</p

Intrinsic membrane properties are unaltered by SAHA treatment.

<p>(A) Representative traces from vehicle (black) and SAHA (red) treated slices during a series of hyperpolarizing and depolarizing current injection steps (scale bar represents 20 mV and 100 ms). There was no difference between vehicle and SAHA treated slices in the number of action potentials elicited by 500 ms current injection pulses at any of the current injection levels (p>0.05, n = 7,7). (B) Action potential threshold, input resistance, and membrane sag reflecting the hyperpolarization-induced inward current, were all unaltered following SAHA treatment (p>0.05). Data are plotted as mean ±SEM.</p

Activity, vocalizations and marking in the urine-exposure open field.

<p>Female-experienced male mice are first habituated to an arena and then presented with a small quantity of urine from a female in estrous. A: During the baseline session 16p11.2 df/+ mice explored the center of the arena more than the WT mice, whereas Cntnap2 -/- showed no differences from its WT control; B: during the exposure session all groups explored the center similarly; C: Ultrasonic vocalizations emitted during the exposure session showed deficits in the Cntnap2 -/- mice but not in the 16p11.2 df/+ as compared to their WT controls; D: Similarly, Cntnap2 -/-, but not 16p11.2 df/+ mice, showed deficits in scent marking during both baseline and urine exposure. Data shown are means ± SEM. (*p < .05; **p < .01).</p

The reciprocal social interaction test did not show deficits in social behavior in either model.

<p>A: The time in which the mice were closer than 5 cm was similar for the two mutants as compared to their respective WT controls. All behaviors with the exception of the active, passive and reciprocal interactions were recorded using an automated system. B: During social interactions, 16p11.2 df/+ pairs were closer to each other as compared to the WT control pairs, whereas Cntnap2 -/- pairs did not show a difference compared to their corresponding WT control pairs; C: Number of interactions of the nose of a mouse towards the front, side or back of the paired mouse. 16p11.2 df/+ mice showed a slight increase in the number of interactions, but only for the interactions towards the back. Cntnap2 -/- mice were similar to their control WT mice; D: Active, passive and reciprocal interactions were similar between mutants and their respective controls. Data shown are means ± SEM. (*p < .05).</p

The Geotaxis Test for the Two Models at the Three Ages Studied.

<p>Mice were placed on an inclined plane and their behavior was observed. A fall indicates lack of coordination whereas a turn indicates a geotactic reflex response. After P7 this reflex is lost and other behaviors appear, such as walking down the plane or sideways.</p><p>The Geotaxis Test for the Two Models at the Three Ages Studied.</p