BackgroundAlzheimer’s disease (AD) is a multifactorial disorder associated with the accumulation of soluble forms of beta-amyloid (Aβ) and its subsequent deposition into plaques. One of the major contributors to neuronal death is chronic and uncontrolled inflammatory activation of microglial cells around the plaques and their secretion of neurotoxic molecules. A shift in microglial activation towards a phagocytic phenotype has been proposed to confer benefit in models of AD. Peroxisome proliferator activator receptor δ (PPARδ) is a transcription factor with potent anti-inflammatory activation properties and PPARδ agonism leads to reduction in brain Aβ levels in 5XFAD mice. This study was carried out to elucidate the involvement of microglial activation in the PPARδ-mediated reduction of Aβ burden and subsequent outcome to neuronal survival in a 5XFAD mouse model of AD.Methods5XFAD mice were orally treated with the PPARδ agonist GW0742 for 2 weeks. The brain Aβ load, glial activation, and neuronal survival were assessed by immunohistochemistry and quantitative PCR. In addition, the ability of GW0742 to prevent direct neuronal death as well as inflammation-induced neuron death was analyzed in vitro.ResultsOur results show for the first time that a short treatment period of 5XFAD mice was effective in reducing the parenchymal Aβ load without affecting the levels of intraneuronal Aβ. This was concomitant with a decrease in overall microglial activation and reduction in proinflammatory mediators. Instead, microglial immunoreactivity around Aβ deposits was increased. Importantly, the reduction in the proinflammatory milieu elicited by GW0742 treatment resulted in attenuation of neuronal loss in vivo in the subiculum of 5XFAD mice. In addition, whereas GW0742 failed to protect primary neurons against glutamate-induced cell death, it prevented inflammation-induced neuronal death in microglia-neuron co-cultures in vitro.ConclusionsThis study demonstrates that GW0742 treatment has a prominent anti-inflammatory effect in 5XFAD mice and suggests that PPARδ agonists may have therapeutic utility in treating AD.

Formation of long-range axons occurs over multiple stages of morphological maturation. However, the intrinsic transcriptional mechanisms that temporally control different stages of axon projection development are unknown. Here, we addressed this question by studying the formation of mouse serotonin (5-HT) axons, the exemplar of long-range profusely arborized axon architectures. We report that LIM homeodomain factor 1b (Lmx1b)-deficient 5-HT neurons fail to generate axonal projections to the forebrain and spinal cord. Stage-specific targeting demonstrates that Lmx1b is required at successive stages to control 5-HT axon primary outgrowth, selective routing, and terminal arborization. We show a Lmx1b→Pet1 regulatory cascade is temporally required for 5-HT arborization and upregulation of the 5-HT axon arborization gene, Protocadherin-alphac2, during postnatal development of forebrain 5-HT axons. Our findings identify a temporal regulatory mechanism in which a single continuously expressed transcription factor functions at successive stages to orchestrate the progressive development of long-range axon architectures enabling expansive neuromodulation.

Serotonin (5-HT) is a crucial neuromodulator linked to many psychiatric disorders. However, after more than 60 years of study, its role in behavior remains poorly understood, in part because of a lack of methods to target 5-HT synthesis specifically in the adult brain. Here, we have developed a genetic approach that reproducibly achieves near-complete elimination of 5-HT synthesis from the adult ascending 5-HT system by stereotaxic injection of an adeno-associated virus expressing Cre recombinase (AAV-Cre) into the midbrain/pons of mice carrying a loxP-conditional tryptophan hydroxylase 2 (Tph2) allele. We investigated the behavioral effects of deficient brain 5-HT synthesis and discovered a unique composite phenotype. Surprisingly, adult 5-HT deficiency did not affect anxiety-like behavior, but resulted in a robust hyperactivity phenotype in novel and home cage environments. Moreover, loss of 5-HT led to an altered pattern of circadian behavior characterized by an advance in the onset and a delay in the offset of daily activity, thus revealing a requirement for adult 5-HT in the control of daily activity patterns. Notably, after normalizing for hyperactivity, we found that the normal prolonged break in nocturnal activity (siesta), a period of rapid eye movement (REM) and non-REM sleep, was absent in all animals in which 5-HT deficiency was verified. Our findings identify adult 5-HT as a requirement for siestas, implicate adult 5-HT in sleep-wake homeostasis, and highlight the importance of our adult-specific 5-HT-synthesis-targeting approach in understanding 5-HT's role in controlling behavior.

Coordinated serotonin (5-HT) synthesis and reuptake depends on coexpression of Tph2, Aadc (Ddc) and Sert (Slc6a4) in brain 5-HT neurons. However, other gene products play critical roles in brain 5-HT synthesis and transport. For example, 5-HT synthesis depends on coexpression of genes encoding the enzymatic machinery necessary for the production and regeneration of tetrahydrobiopterin (BH4). In addition, the organic cation transporter 3 (Oct3, Slc22a3) functions as a low affinity, high capacity 5-HT reuptake protein in 5-HT neurons. The regulatory strategies controlling BH4 and Oct3 gene expression in 5-HT neurons have not been investigated. Our previous studies showed that Pet-1 is a critical transcription factor in a regulatory program that controls coexpression of Tph2, Aadc and Sert in 5-HT neurons. Here, we investigate whether a common regulatory program determines global 5-HT synthesis and reuptake through coordinate transcriptional control. We show with comparative microarray profiling of flow sorted YFP+ Pet-1−/− and wild type 5-HT neurons that Pet-1 regulates BH4 pathway genes, Gch1, Gchfr, and Qdpr. Thus, Pet-1 coordinates expression of all rate-limiting enzymatic (Tph2, Gch1) and post-translational regulatory (Gchfr) steps that determine the level of mammalian brain 5-HT synthesis. Moreover, Pet-1 globally controls acquisition of 5-HT reuptake in dorsal raphe 5-HT neurons by coordinating expression of Slc6a4 and Slc22a3. In situ hybridizations revealed that virtually all 5-HT neurons in the dorsal raphe depend on Pet-1 for Slc22a3 expression; similar results were obtained for Htr1a. Therefore, few if any 5-HT neurons in the dorsal raphe are resistant to loss of Pet-1 for their full neuron-type identity.

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