Cyclin-dependent kinase 5 (Cdk5) is a proline-directed serine/threonine kinase. We have previously reported that Cdk5 participates in the regulation of nociceptive signaling, and the expression of Cdk5 and its activator, p35, are up-regulated in nociceptive neurons during peripheral inflammation. The aim of our current study was to identify the proinflammatory molecules that regulate Cdk5/p35 activity in response to inflammation. We constructed a vector that contains the mouse p35 promoter driving luciferase expression. We transiently transfected this vector in PC12 cells to test the effect of several cytokines on p35 transcriptional activity and Cdk5 activity. Our results indicate that tumor necrosis factor-␣ (TNF-␣) activates p35 promoter activity in a dose-and time-dependent manner and concomitantly up-regulates Cdk5 activity. Because TNF-␣ is known to activate ERK1/2, p38 MAPK, JNK, and NF-B signaling pathways, we examined their involvement in the activation of p35 promoter activity. MEK inhibitor, which inhibits ERK activation, decreased p35 promoter activity, whereas the inhibitors of p38 MAPK, JNK, and NF-〉 increased p35 promoter activity, indicating that these pathways regulate p35 expression differently. The mRNA and protein levels of Egr-1, a transcription factor, were increased by TNF-␣ treatment, and this increase was dependent on ERK signaling. In a mouse model of inflammation-induced pain in which carrageenan injection into the hind paw causes hypersensitivity to heat stimuli, TNF-␣ mRNA was increased at the site of injection. These findings suggest that TNF-␣-mediated regulation of Cdk5 activity plays an important role in inflammation-induced pain signaling.

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Prenatal stress causes predisposition to cognitive and emotional disturbances and is a risk factor towards the development of neuropsychiatric conditions like depression, bipolar disorders and schizophrenia. The extracellular protein Reelin, expressed by Cajal-Retzius cells during cortical development, plays critical roles on cortical lamination and synaptic maturation, and its deregulation has been associated with maladaptive conditions. In the present study, we address the effect of prenatal restraint stress (PNS) upon Reelin expression and signaling in pregnant rats during the last 10 days of pregnancy. Animals from one group, including control and PNS exposed fetuses, were sacrificed and analyzed using immunohistochemical, biochemical, cell biology and molecular biology approaches. We scored changes in the expression of Reelin, its signaling pathway and in the methylation of its promoter. A second group included control and PNS exposed animals maintained until young adulthood for behavioral studies. Using the optical dissector, we show decreased numbers of Reelin-positive neurons in cortical layer I of PNS exposed animals. In addition, neurons from PNS exposed animals display decreased Reelin expression that is paralleled by changes in components of the Reelin-signaling cascade, both in vivo and in vitro. Furthermore, PNS induced changes in the DNA methylation levels of the Reelin promoter in culture and in histological samples. PNS adult rats display excessive spontaneous locomotor activity, high anxiety levels and problems of learning and memory consolidation. No significant visuo-spatial memory impairment was detected on the Morris water maze. These results highlight the effects of prenatal stress on the Cajal-Retzius neuronal population, and the persistence of behavioral consequences using this treatment in adults, thereby supporting a relevant role of PNS in the genesis of neuropsychiatric diseases. We also propose an in vitro model that can yield new insights on the molecular mechanisms behind the effects of prenatal stress.

Injury and inflammation trigger activation of several critical cellular pathways in nociceptive signaling in the peripheral nervous system, but their precise molecular mechanisms have not been clearly defined. Cyclin-dependent kinase 5 (Cdk5), a serine/threonine kinase, is mainly expressed in the post-mitotic neurons, and has many important roles in the development, functions and pathophysiology of diseases of the nervous system. Although many functional roles of Cdk5 have been identified in neurons, its precise role in pain signaling has not been well determined. Experimental inflammation in the hind paws of mice resulted in increased mRNA and protein levels of Cdk5 and its activator p35, as well as the Cdk5 activity in nociceptive neurons (Pareek et al., 2006). Furthermore, we also identified that Cdk5 phosphorylates transient receptor potential vanilloid 1 (TRPV1), a key receptor that modulates agonist-induced calcium influx in the neurons (Pareek et al., 2007). We subsequently demonstrated that inflammation triggers increase in Cdk5 activity through activation of early growth response 1 (Egr-1) and p35 expression by tumor necrosis factor alpha (TNF-α) (Utreras et al., 2009). These findings suggest that Cdk5 plays an important role in pain signaling and therefore Cdk5 and its activators are potentially important drug targets for development of novel analgesics to treat neuropathic pain.