Recognition memory and anxiety like behavior is affected by GluA2 aAB

our passive-transfer animal models. As behavioral readouts we used the novel object recognition test as a method for memory testing and the elevated plus maze test (EPM) or the black and white box as methods to assess anxiety related behavior. Additionally, we used an infrared (IR) actimeter for the analysis of locomotor activity. With this set of behavioral experiments we were able to reliably recognize behavioral changes of mice after aAB administration.

The IR actimeter is measuring motoric activity, including horizontal activity, local motor activity and rearings. Data from the IR actimeter were used to detect influence of the surgery on the activity of the animals. Moreover, this data can also be used to assess the direct influence of aAB administration on motoric behavior (Caille et al. 1999, Berrendero et al. 2005). As we expected, we did not find changes in locomotor activity since motor dysfunction is not a key symptom of the disease. This result also allows us to conclude that the surgeries had no influence on overall activity of test animals in the animal models (Planaguma et al. 2016, Haselmann et al. in revision).

The anxiety tests both measure the mice’s natural aversion to open spaces and the tendency to be thigmotaxic. In the EPM there are two closed arms allowing the animal to hide in a dark zone and two open arms that are enlightened and allow the view over a wide area of the room. Dependent on anxiety level of the animal it will remain in the dark for a longer period (Walf and Frye 2007, Pellow et al. 1985). In the black and white box, the same paradigm is tested by using a black darkened box as a possibility to hide and an enlightened white box as a stimulus for anxiety (Gimenez-Llort et al. 2015). In contrast to the assumption that anxiety is nearly exclusively mediated by the amygdala, recent data revealed striking evidence that the ventral hippocampus is involved, too (Bannerman et al. 2014, Bannerman et al. 2004). In our experiments we could show that GluA2 aABs mediate increased anxiety in the hippocampal injection animal model in the EPM, as well as in the ventricular infusion animal model in the black and white box (Haselmann et al. in revision). Whereas the osmotic pump model could account for aAB induced pathomechanisms in the hippocampus as well as in the amygdala, the injection model is likely to induce anxiety related behavior especially in the hippocampus since the diffusion of locally injected aABs into the amygdala is not expected. Together, these findings are in line with further studies suggesting increased anxiety-like behavior after

146 5 Discussion benzodiazepine withdrawal induced deregulation of synaptic AMPAR composition (Van Sickle et al. 2004, Das et al. 2008).

We used the Novel Object Recognition (NOR) test in order to test the impact of anti-AMPAR aABs on memory. This test uses the animals innate exploratory behavior without the need of other positive or negative stimuli (Antunes and Biala 2012). Another advantage of the NOR test is the possibility that it can be repeated several times with different objects without disturbing the performance of the animal. This allows the experimenter to test recognition memory before and after surgery to rule out effects of the surgery on brain function. The test measures the discrimination index, e.g. the time spent exploring the unknown object divided by the total time exploring both objects. A higher discrimination index is considered to reflect greater memory retention for the familiar object (Ennaceur and Delacour 1988). There is still discussion about the participating brain regions in NOR. On the one side, involvement of different brain regions in novel object preference and object location paradigms is suggested (Barker and Warburton 2011). Novel object preference may be mediated more by perirhinal cortex, whereas object location is mediated by the hippocampus. On the other side, also the hippocampus is identified to participate in novel object preference test (Broadbent et al. 2010, Cohen and Stackman 2015). Overall, the involvement of the hippocampus in object recognition memory seems to be dependent on the delay between the familiarization and the test phase (Cohen and Stackman 2015, Hammond et al. 2004). In our experiments we used two objects on a diagonal axis of the open field box for habituation and exchanged one of these objects for the acquisition (Puighermanal et al. 2009). Our experiments revealed a decrease in object recognition memory by GluA2 aABs in the microinjection model as well as the osmotic pump model. This reflects the characteristic anterograde memory deficit that is characteristic in patients with GluA2 AE (Tuzun and Dalmau 2007).

Of note, in the parenchymal injection model of GluA2 AE also control IgG injected mice showed slightly decreased performance in the NOR task in comparison to the baseline performance before surgery. This effect could be due to repetitive surgeries and anesthesia. Furthermore, repetitive injections at multiple injection sites might have effects on behavior due to small lesions of brain tissue. Another possible influence factor might be the short time (one day) of recovery after the last surgery. However, the decrease in recognition memory of these control mice was by far less pronounced than in GluA2 IgG injected animals. Based on these caveats the osmotic pump model might be better suited for behavioral testing whereas the injection model is possibly more suitable for electrophysiological recordings due to its

defined local application and the exact amount of injected aAB fractions to these targeted injection sites.

5.6 The effects of aABs to ionotropic glutamate receptors on synaptic long-term