Gabriela Paglini

The Cdk5‐p35 kinase associates with the Golgi apparatus and regulates membrane traffic

We show here that an active Cdk5‐p35 kinase is present in Golgi membranes, where it associates with a detergent‐insoluble fraction containing actin. In addition, Cdk5‐p35‐dependent phosphorylation of α‐PAK immunoreactive protein species was detected in Golgi membranes, as well as an interaction with the small GTPase, Cdc42. Moreover, antisense oligonucleotide suppression of Cdk5 or p35 in young cultured neurons, as well as inhibition of Cdk5 activity with olomoucine, blocks the formation of membrane vesicles from the Golgi apparatus. Taken together, these results show a novel subcellular localization of this kinase and suggest a role for Cdk5‐p35 in membrane traffic during neuronal process outgrowth.

Tau Protein Function in Axonal Formation

Tau protein is a predominantly neuronal microtubule-associated protein that is enriched in axons and is capable of promoting microtubule assembly and stabilization. In the present article we review some of the key experiments directed to obtain insights about tau protein function in developing neurons. Aspects related to whether or not tau has essential, unique, or complementary functions during axonal formation are discussed.

Involvement of septal Cdk5 in the emergence of excessive anxiety induced by stress

The aim of the present study was to evaluate whether the activation of Cdk5, a protein that has been suggested to participate in higher cognitive functions, is required for the onset of a sensitized anxiety-related behavior induced by stress. The exposure to restraint enhanced both Cdk5 expression in certain subareas of the septohippocampal system, principally in the lateral septum (LS) and septal Cdk5 kinase activity in rats. Behaviorally, restrained wild type mice showed a behavior indicative of enhanced anxiety in the elevated plus maze (EPM). In contrast, unstressed mice and stressed knockout mice, which lacked the p35 protein, the natural activator of Cdk5, displayed similar anxiety-like behavior in the EPM. Finally, the intra-LS infusion of olomoucine - a Cdk5 inhibitor - blocked the enhanced anxiety in the EPM induced by prior stress in rats. All these data provide evidence that septal Cdk5 is required in the emergence of a sensitized emotional process induced by stress.

Mice lacking p35 display hyperactivity and paradoxical response to psychostimulants

J. Neurochem. (2010) 114, 203–214.

Previous stress exposure enhances both anxiety-like behaviour and p35 levels in the basolateral amygdala complex: Modulation by midazolam

Stress exposure induces long lasting neurobiological changes in selected brain areas, which could be associated with the emergence of negative emotional responses. In the present study, previously restrained animals exhibited excessive anxiety one day later in the elevated plus maze. We explore whether stress exposure affects the expression levels of cyclin-dependent kinase 5 (Cdk5) and of its activator protein p35, in diverse amygdaloid nuclei. Stress exposure enhanced p35 levels selectively in the basolateral amygdala (BLA). This up-regulation might be functionally associated with the occurrence of exaggerated anxiety since such emotional response was selectively reversed by an intra-BLA infusion of olomoucine, a Cdk5 inhibitor, 15 min prior to the restraint session. Moreover, pre-treatment with midazolam, a benzodiazepine ligand, not only prevented the excessive anxiety but also attenuated the p35 increase in the BLA of stressed rats. In conclusion, we suggest a pivotal role of the Cdk5/p35 complex, specifically in BLA in the excessive anxiety induced by a previous stressful experience.

Transient Enhanced Expression of Cdk5 Activator p25 after Acute and Chronic d-Amphetamine Administration

The cellular and molecular mechanisms of sensitization in the addictive process are still unclear. Recently, chronic treatment with cocaine has been shown to upregulate the expression of cyclin‐dependent kinase 5 (cdk5) and its specific activator, p35, in the striatum, as a downstream target gene of ΔFosB, and has been implicated in compensatory adaptive changes associated with psychostimulants. Cdk5 is a serine/threonine kinase and its activation is achieved through association with a regulatory subunit, either p35 or p39. P35 is cleaved by the protease calpain, which results in the generation of a truncated product termed p25, which contains all elements necessary for cdk5 activation. The cdk5/p35 complex plays an essential role in neuronal development and survival. It has also been involved in neuronal trafficking and transport and in dopaminergic transmission, indicating its role either in presynaptic and postsynaptic signaling. In this study we report that the cdk5/p35 complex participates in acute and chronic d‐amphetamine (AMPH)‐evoked behavioral events, and we show a surprisingly transient enhanced expression of p25 and a lasting increased expression of p35 in dorsal striatal synaptosomes after acute and chronic AMPH administration. Pak1, a substrate for cdk5, is also enriched in the synaptosomal fraction of acute AMPH‐treated rats. Our data suggest that the transient upregulation of p25 may regulate the activity of cdk5 in phosphorylating particular substrates, such as Pak1, implicated in the compensatory adaptive morphophysiologic changes associated with the process of behavioral sensitization to psychostimulants.

Association between the expression of amphetamine-induced behavioral sensitization and Cdk5/p35 activity in dorsal striatum.

Sensitization to psychostimulants is strongly influenced by the environmental context in which the drug is administered and little is known about the molecular mechanisms that regulate this process. Chronic treatment with psychostimulants has been shown to upregulate the expression of cyclin-dependent kinase 5 (Cdk5) in the striatum, as a downstream target gene of ΔFosB. This study was therefore designed to analyze neurochemical changes underlying the expression of amphetamine-induced sensitization. To this end, periadolescent rats were given saline or 4 mg/kg amphetamine in a NOVEL or a HOME environment. After 1 day, subjects were challenged with vehicle or 2 mg/kg amphetamine in the same context in which they received the first administration of the drug. Locomotor activity and the expression levels of p35 and Cdk5 activity in synaptosomes of the dorsal striatum were analyzed. The expression of behavioral sensitization was observed only in the NOVEL condition. Furthermore, only animals trained and tested in the NOVEL condition showed increased p35 protein levels and Cdk5 activity. Our findings provide clear behavioral and neurochemical evidence of a specific association between increased p35 and Cdk5 activity in the dorsal striatum and the expression of amphetamine-behavioral sensitization, allowing us to propose p35 as a biochemical marker of behavioral sensitization to amphetamine.

Role of the Golgi Apparatus During Axon Formation

During recent years, a large body of evidence has accumulated regarding cellular events and potential mechanisms underlying the development of specialized cellular projections, namely axons and dendrites in mammalian neurons. Most of these studies have focused on the regulation of cytoskeletal organization and function during different stages of neuronal polarization. Much less effort has been invested in analyzing how cytoskeletal assembly and dynamics are coordinately coupled with membrane addition at sites of active growth or with membrane formation and/or exit from the Golgi complex. In this chapter, we shall review current evidence concerning the participation of the Golgi apparatus in axon specification, one of the earliest events in neuronal polarization.