They were maintained on a 12/12-h light/dark cycle and fed administration of drugs that affect DA release result in changes in PKC activity in the striatum (Giambalvo, 1988; Giambalvo, 1989). TH at ser 40. Therefore, these results suggest that the MA-induced enhancement of PKC expression is a critical factor in the impairment of TH phosphorylation at ser 40 and that pharmacological or genetic inhibition of PKC may be protective against MA-induced dopaminergic neurotoxicity (Dunkley et al., 2004; Hufton et al., 1995). Of the phosphorylation sites at the N-terminus of TH only ser 31 and ser 40 are readily phosphorylated and activate TH (Haycock and Wakade, 1992; Sutherland et al., 1993). The protein kinase C (PKC) family consists of serine/threonine kinases and is broadly classified into three subgroups based on sensitivity to important cofactors, including phospholipids and Ca2+ (Dempsey et al., 2000; Gschwendt, 1999). The conventional PKC isoforms (, I, II, ) are sensitive to Ca2+ and diacylglycerol and the novel isoforms (, , , , ) are Ca2+ impartial but require diacylglycerol for activation. The atypical isoforms (, /) require neither Ca2+ nor diacylglycerol for activation. PKC isoforms are differentially distributed in tissues and play key roles in various cellular biological processes, including cell differentiation and growth, apoptosis, tumor suppression, and carcinogenesis. In most studies, PKC inhibitors are used to demonstrate the anti-apoptotic role of the PKC family. Of the novel isoforms, PKC was the first member found to be functionally modulated by tyrosine phosphorylation upon H2O2 treatment (Konishi et al., 1997; Steinberg, 2004). A number of studies have found that the proteolytic activation of PKC plays a key role in apoptotic cell death of dopaminergic neurons (Kaul et al., 2003; Yang et al., 2004; Kitazawa et al., 2003; Latchoumycandane et al., 2005; Kanthasamy et al., 2006). However, little is known concerning the role of PKC during dopaminergic toxicity induced by an amphetamine analog. Thus, the involvement of PKC in methamphetamine (MA)-induced dopaminergic toxicity is usually examined here. It was observed that PKC is usually critically involved in MA-induced dopaminergic toxicity and that PKC inhibition using the PKC inhibitor rottlerin or a PKC gene knockout (?/?) mouse model attenuates MA-induced dopaminergic toxicity through the upregulation of TH phosphorylation at ser 40. As recent reports indicate that rottlerin-mediated pharmacological effects as a PKC inhibitor are somewhat controversial (Soltoff, 2007; IACS-9571 Susarla et al., 2003; Tapia et al., 2006), an additional experiment using a PKC IACS-9571 antisense oligonucleotide was performed. Material and Methods Animals All mice were treated in accordance IACS-9571 with the NIH Guideline for the Humane Care and Use of Laboratory FIGF Animals. They were maintained on a 12/12-h light/dark cycle and fed IACS-9571 administration of drugs that affect DA release result in changes in PKC activity in the striatum (Giambalvo, 1988; Giambalvo, 1989). Thus, the striatal expression of PKC after the final MA dose was examined (Fig.3). Some PKC expression was observed in the absence of MA in PKC (+/+) mice although treatment with MA significantly increased PKC expression ((Campbell et al., 1986; Wu et al., 1992). Zhang et al. (2007a) found a high expression of PKC in dopaminergic neurons and initially hypothesized that PKC might phosphorylate TH to increase its activity. To test this, they used the PKC inhibitor rottlerin to inhibit the kinase and anticipated that inhibition of PKC would result in inhibition of TH activity. Unexpectedly, a dose-dependent increase in TH activity and DA IACS-9571 levels was observed in cells treated with rottlerin. Similar to the current data, Zhang et al. (2007b) provided evidence that rottlerin treatment can rescue TH-positive neurons from MPP+-induced neurotoxicity model to study the death of dopaminergic neurons (Takahashi et al., 1994; Tian et al., 2007; Wang et al., 2008; Suwanjang et al., 2010; Tiong et al., 2010). The PKC family consists of at least 12 isozymes of which PKC and PKC are expressed in SH-SY5Y dopaminergic neuroblastoma cells (Zeidman et al., 1999; Mackay and Mochly-Rosen, 2001; Pan et al., 2008). However, inhibition of PKC with rottlerin did not reverse the cell injury caused by 6-OHDA in SHSY5Y cells (Tiong et al., 2010). Although MA treatment significantly reduced the viability of SH-SY5Y cells in a concentration-related manner in our pilot study, rottlerin (at a level of 5M) did not significantly affect MA-induced reduced viability of SH-SY5Y cells (data not shown). Similarly, PC12 pheochromocytoma cells have been widely used to study the molecular.