Behavioural Brain Research 280 (2015) 45–50 Contents lists available at ScienceDirect Behavioural Brain Research jou rn al hom epage: www.elsevier.com/locate/bbr Research report Networks of protein kinases and phosphatases in the individual phases of contextual fear conditioning in the C57BL/6J mouse Goran Mucic a , Sunetra Sase a , Oliver Stork b , Gert Lubec a,∗ , Lin Li a,∗ a Department of Pediatrics, Medical University of Vienna, 1090 Vienna, Austria b Institute of Biology, Otto von Guericke University, Magdeburg, 39120 Magdeburg, Germany h i g h l i g h t s • Protein kinases and phosphatases differ between individual phases of fear conditioning. • Protein arrays were used to screen different signalling protein levels between phases. • Protein phosphatase 2A was linked to the retrieval phase of cFC. a r t i c l e i n f o Article history: Received 1 November 2014 Received in revised form 8 November 2014 Accepted 12 November 2014 Available online 21 November 2014 Keywords: Contextual fear conditioning Protein kinase Kinase network Protein phosphatase a b s t r a c t Although protein kinases and phosphatases have been reported to be involved in fear memory, infor- mation about these signalling molecules in the individual phases of contextual fear conditioning (cFC) is limited. C57BL/6J mice were tested in cFC, sacrificed and hippocampi were used for screening of approximately 800 protein kinases and phosphatases by protein microarrays with subsequent Western blot confirmation of threefold higher or lower hippocampal levels as compared to foot shock controls. Immunoblotting of the protein kinases and phosphatases screened out was carried out by Western blot- ting. A network of protein kinases and phosphatases was generated (STRING 9.1). Animals learned the task in the paradigm and protein kinase and phosphatase levels were determined in the individual phases acquisition, consolidation and retrieval and compared to foot shock controls. Protein kinases discoidin containing receptor 2 (DDR2), mitogen activated protein kinase kinase kinase 7 (TAK1), pro- tein phosphatases dual specificity protein phosphatase (PTEN) and protein phosphatase 2a (PP2A) were modulated in the individual phases of cFC. Phosphatidyl-inositol-3,4,5-triphosphate 3-phosphatase and phosphatidylinositol-3 kinase (PI3K) that is interacting with PTEN were modulated as well. Freezing time was correlating with PP2A levels in the retrieval phase of cFC. The abovementioned protein kinases, phos- phatases and inositol-signalling enzymes were not reported so far in cFC and the results are relevant for interpretation of previous and design of future studies in cFC or fear memory. Protein phosphatase PP2A was, however, the only signalling compound tested that was directly linked to retrieval in the cFC. © 2014 Published by Elsevier B.V. 1. Introduction Although protein kinases are key elements in memory mech- anisms, published data on hippocampal protein kinases and protein phosphatases in contextual fear conditioning (cFC) is limited: It has been shown that conditional deletion of the erk 5 gene results in impaired contextual fear memory consolidation but not in acquisition and retrieval. Moreover, mitogen-activated protein ∗ Corresponding authors. Tel.: +43 1 40400 3215; fax: +43 1 40400 6065. E-mail addresses: gert.lubec@meduniwien.ac.at (G. Lubec), linli@gmail.com (L. Li). kinase 7 (Erk5 MAP) may play a role in the establishment of remote contextual fear memory [1]. Atkins et al. have shown that hippocampal phosphorylated mitogen-activated protein kinase 1 (MAPK 1) but not mitogen-activated protein kinase 3 (MAPK3) increased one hour following cFC. In addition, the authors have shown that inhibition of dual specificity mitogen-activated pro- tein kinase kinase 1(MEK), which is responsible for the activation of mitogen-activated protein kinase (MAPK), blocks associative learning in fear conditioning. Many protein kinases such as MAPK, protein kinase C (PKC) and calcium/calmodulin-dependent protein kinase type II subunit alpha (-CaMKII) are activated in the hip- pocampus after fear conditioning, however, not at the same time [2] and the role of this protein kinase for the formation of a variety of fear memories was described [3]. http://dx.doi.org/10.1016/j.bbr.2014.11.024 0166-4328/© 2014 Published by Elsevier B.V.