L., 2002). Offered the reduced anxiousness in Rcan1 KO mice, we tested these mice for abnormal PPI. We located no difference in PPI in between Rcan1 KO mice and WT littermates across a selection of acoustic prepulse intensities (Fig. 4D; percentage inhibition of startle response: 74 dB, t(26) 0.123, p 0.9; 78 dB, t(26) 0.601, p 0.5; 82 dB, t(26) 1.232, p 0.two; 86 dB, t(26) 1.222, p 0.two; 90 dB, t(26) 1.753, p 0.091; startle test: t(26) 0.113, p 0.9; null period: t(26) 0.109, p 0.9). This demonstrates that the anxiety phenotype in Rcan1 KO mice isn’t the outcome of abnormal sensorimotor gating. Considering the fact that RCAN1 removal decreased the show of anxiety in Rcan1 KO mice, we next tested whether RCAN1 overexpression could enhance anxiousness behaviors. We took benefit of two conditional flox-ON RCAN1 transgenic mouse lines (Tg1 and Tg1a) that overexpress human RCAN1 protein at higher or low levels, respectively, inside the presence of Cre recombinase (Oh et al., 2005). We utilized two Cre-driver lines to activate RCAN1 overexpression at distinct developmental time points, Nse-Cre in the course of D2 Receptor Agonist medchemexpress development (onset at about embryonic day 16.5; Forss-Petter et al., 1990) and T29-Bradykinin B2 Receptor (B2R) Antagonist Storage & Stability CamkII -Cre postdevelopmentally (onset at about postnatal day 14; Hoeffer et al., 2008). Overexpression of RCAN1 was confirmed by Western blots making use of antibodies against RCAN1 (Vega et al., 2003; Hoeffer et al., 2007) along with the FLAG epitope tagged for the RCAN1 transgenic construct (Oh et al., 2005; Fig. 4E). RCAN1 overexpression applying either Cre driver had no detectable impact in the OFA assay (Table 1). Within the EPM assay, nevertheless, RCAN1 overexpression early in development below Nse-Cre in RCAN1Tg1a mice was shown to lower open-arm time compared with control WT (no Cre) littermates (Mann?Whitney U(83) two.010, p 0.044; Fig. 4F ). This impact was not due to group variations in locomotor activity (distance moved t(18) 1.683, p 0.110) or sensorimotor gating (Table two), which supports the concept that the decreased open-arm time in NseRCAN1Tg1a mice represents greater anxiety. Having said that, overexpression of the other RCAN1 construct (RCAN1Tg1) under the exact same Nse-Cre driver did not influence EPM open-arm time, (Mann?Whitney U(18) 0.140, p 0.9; Fig. 4F ). Also, postdevelopmental RCAN1 overexpression under CamkII -Cre didn’t affect EPM open-arm time (CamkII -RCAN1Tg1a open-arm time, Mann hitney U(70) 0.018, p 0.9; CamkII RCAN1Tg1 open-arm time, Mann hitney U(28) 0.873, p 0.four; Fig. 4F ). Combined together with the behavioral final results in16936 ?J. Neurosci., October 23, 2013 ?33(43):16930 ?Hoeffer, Wong et al. ?RCAN1 Modulates Anxiety and Responses to SSRIsADBECFFigure four. Rcan1 KO mice show decreased measures of anxiety inside the EPM. A, Rcan1 KO mice spend significantly more time exploring the open arms in the EPM compared with their WT littermates. N ten KO, 12 WT. B, Rcan1 KO mice enter the open arms early inside the EPM test (minute 1) whereas their WT littermates increased open-arm exploration starting at the third minute of testing compared with minute 1. N ten KO, 9 WT. C, Total distance moved and speed of Rcan1 KO mice are indistinguishable from WT mice inside the EPM. N 10 KO, 12 WT. D, Rcan1 KO mice display related PPI of acoustic startle responses compared with their WT littermates. E, Western blot analysis of RCAN1 expression within the PFC of RCAN1 transgenic (Tg) mice utilised for this study. Upper blot is stained with an RCAN1 antibody that recognizes endogenously expressed RCAN1.1L ( 38 kDa) and RCAN1.4 ( 28 kDa) protein isoforms and transgenicall.
