Such segregation of AP-dependent and AP-independent presynaptic regulatory mechanisms may be particularly significant in circumstances where evoked release probability is low (Borst, 2010), and the two types of synaptic signals are difficult to differentiate. Under these circumstances, selective augmentation of spontaneous neurotransmitter release may facilitate neurotrophic, homeostatic PI3K inhibitor or other signaling functions of released neurotransmitter substances without compromising their function in AP-evoked information transfer. Recent studies suggest that the AP-independent forms of neurotransmitter release are critical

in the regulation of behavioral outcomes such as nociception, memory processing, and response to antidepressants (Andresen et al., 2012, Autry et al., 2011, Jin et al., 2012, Kavalali and Monteggia, 2012, Xu et al., 2012 and Nosyreva et al., 2013). This premise is consistent with the recent

behavioral analysis of VAMP7 knockout mice that revealed a deficit in anxiety-related behaviors (Danglot et al., 2012). In this way, identification of the vesicular release machineries and neuromodulators that specifically modify AP-independent forms of neurotransmission may provide novel avenues to manipulate neurotransmission without altering AP-dependent information processing. These types of approaches provide a promising strategy to uncover the functional roles FK228 of these unconventional forms of neuronal communication in the regulation of behavior in normal as well as in disease states. Dissociated hippocampal cultures were prepared from postnatal day 0–3 Sprague-Dawley rats of either sex

as described previously (Kavalali et al., 1999). For syb2 knockout (KO) and SNAP-25 KO experiments, dissociated hippocampal cultures were prepared from embryonic day 18 mice constitutively deficient in syb2 (syb2−/−) or SNAP-25 (SNAP-25−/−) as well as their littermate controls (Schoch et al., 2001 and Washbourne et al., 2002). ApoER2 KO and VLDLR KO cultures were prepared from mice generated by constitutive deletion of Apoer2 ( Trommsdorff et al., 1999) and vldlr genes ( Frykman et al., 1995). To generate below neurons deficient in p110α and p110β isoforms of PI3K (gift of Drs. Joel Elmquist, UT Southwestern, and Jean Zhao, Dana-Farber Cancer Institute), hippocampal cultures from mice expressing conditionals alleles of p110α and p110β genes were infected with lentivirus expressing Cre. Lentiviral expression system shows high infection efficacy (>90%) as previously demonstrated by full rescue of synaptic transmission in syb2−/− cultures by lentiviral expression of syb2 ( Deák et al., 2006). All experiments were performed on 14–21 days in vitro (DIV) cultures. All experiments were performed in accordance with protocols approved by the UT Southwestern Institutional Animal Care and Use Committee.