Interleukin (IL)-12 is a heterodimeric cytokine consisting of the p40 and p35 chains encoded on separate chromosomes. Coordinated expression of the two constituent genes is crucial for appropriate immune responses in timing, location, and magnitude. Interferon (IFN)-γ priming of IL-12 production by macrophages represents an important physiological process in vivo for escalated cellular response to microbial infections. We provide evidence that IFN regulatory factor (IRF)-1–deficient macrophages have a selective impairment in mRNA synthesis of IL-12 p35 but not the p40 gene, and a strong deficiency in the production of IL-12 p70 but not p40. We demonstrate that the levels of IL-12 p35 protein stimulated by IFN-γ and lipopolysaccharide (LPS) correspond to those of its mRNA, and that the nuclear factor κB signaling pathway is essential for the induction of IL-12 p35 transcription by LPS. IRF-1 plays a major role in the transcriptional activation of the IL-12 p35 gene, but not of the p40 gene, by physically interacting with an inverted IRF element within the IL-12 p35 promoter upon IFN-γ activation. Moreover, IRF-1–mediated transcriptional activation of the p35 promoter requires the cooperation of two adjacent Sp1 elements. Thus, IRF-1 acts as a critical component of IFN-γ signaling in the selective activation of IL-12 p35 transcription in synergy with LPS-mediated events.
The in vivo binding of positron emission tomography (PET) and single photon emission computer tomography (SPECT) radiotracers to dopamine D2 receptors in the striatum can be influenced by competition with endogenous dopamine. The present study was undertaken to determine if a similar inhibition of radiotracer binding to dopamine receptors could be observed following pharmacologically-evoked dopamine release in rat brain striatal slices. Striatal slices were incubated in a large volume of oxygenated Krebs saline and exposed to amphetamine or methamphetamine to evoke dopamine release within the slice. Amphetamine and methamphetamine, at concentrations up to 30 microM, reduced [3H]raclopride binding in the slices by 77% and 86%, respectively, with 50% inhibition at 1.6 microM amphetamine or 3.0 microM methamphetamine. Neither drug produced a significant effect on binding of [3H]SCH 23390 in the slices. This suggests that dopamine was able to interfere with radiotracer binding to D2 but not D1 receptors. The dopamine uptake blockers, cocaine and methylphenidate, had relatively little effect by themselves on [3H]raclopride binding but, by inhibiting amphetamine-induced dopamine release, significantly reduced inhibition of [3H]raclopride binding by a low (3 microM) amphetamine concentration. At a higher (30 microM) amphetamine concentration the inhibition of [3H]raclopride binding was not antagonized by uptake blockers and data obtained from homogenate binding experiments indicated a direct displacement of [3H]raclopride binding by amphetamine at this concentration. In conclusion the data obtained in the present study demonstrate that the effects of amphetamine on striatal radiotracer accumulation observed in PET and SPECT can also be observed in brain slices in vitro and, at least at low amphetamine concentrations, are mediated by competition with released dopamine.
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