Dopamine and glutamate induce distinct striatal splice forms of Ania-6, an RNA polymerase II-associated cyclin.

Control of neuronal gene expression by drugs or neurotransmitters is a critical step in long-term neural plasticity. Here, we show that a gene induced in the striatum by cocaine or direct dopamine stimulation, ania-6, is a member of a novel family of cyclins with homology to cyclins K/T/H/C. Further, different types of neurotransmitter stimulation cause selective induction of distinct ania-6 isoforms, through alternative splicing. The longer Ania-6 protein colocalizes with nuclear speckles and is associated with key elements of the RNA elongation/processing complex, including the hyperphosphorylated form of RNA polymerase II, the splicing factor SC-35, and the p110 PITSLRE cyclin-dependent kinase. Distinct types of neuronal stimulation may therefore differentially modulate nuclear RNA processing, through altered transcription and splicing of ania-6.

Effect of a functional impairment of corticostriatal transmission on cortically evoked expression of c-Fos and zif 268 in the rat basal ganglia.

The activity-dependent induction of immediate-early genes is commonly used to map activated neuronal networks. In a previous analysis of the cortico-basal ganglia circuits, we have shown that a cortical stimulation produces Fos protein expression in the striatum and the subthalamic nucleus, with a pattern which conforms to the anatomical organization of cortical projections [Sgambato V. et al. (1996) Neuroscience 81, 93-112]. In the present study, we examined the effects of a unilateral blockade of the corticostriatal transmission on c-fos and zif 268 messenger RNA expression evoked in the substantia nigra pars reticulata and the subthalamic nucleus following stimulation of the ipsilateral motor cortex. The blockade of the corticostriatal pathway was performed either by an excitotoxic striatal lesion or by an application of the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione within the striatum. After application of the glutamate receptor antagonist, which prevented the cortical stimulation activating the GABAergic striatonigral pathway, the induction of both c-fos and zif 268 messenger RNAs was facilitated in the ipsilateral substantia nigra pars reticulata. In the subthalamic nucleus ipsilateral to the application of 6-cyano7-nitroquinoxaline-2,3-dione, the cellular discharges evoked by stimulation of the cortex were considerably shortened as a result of the blockade of the disinhibitory striato-pallido-subthalamic circuit. However, a strong expression of immediate-early genes was still induced by the cortical stimulation. By contrast, after unilateral kainate lesion of the striatum, the cortical stimulation was no longer able to induce c-fos and zif 268 messenger RNA expression in the ipsilateral subthalamic nucleus and in the substantia nigra pars reticulata bilaterally. The lack of immediate-early gene induction strongly contrasted with the neuronal discharges evoked in these nuclei by the cortical stimulation. Comparison between the cortically evoked neuronal activities and the pattern of immediate-early gene expression suggests that the induction of immediate-early genes in the basal ganglia mainly reflects the level of synaptic activity rather than the frequency of discharge of the postsynaptic neurons. Moreover, the results stress that modifications of immediate-early gene expression observed in the basal ganglia after an acute or a chronic interruption of the corticostriatal transmission are not superimposable.

Extracellular signal-regulated kinase (ERK) controls immediate early gene induction on corticostriatal stimulation.

Activity-dependent changes in neuronal structure and synaptic remodeling depend critically on gene regulation. In an attempt to understand how glutamate receptor stimulation at the membrane leads to gene regulation in the nucleus, we traced intracellular signaling pathways targeting DNA regulatory elements of immediate early genes (IEGs). For this purpose we used an in vivo electrical stimulation of the glutamatergic corticostriatal pathway. We show that a transient activation of extracellular signal-regulated kinase (ERK) proteins (detected by immunocytochemistry with an anti-active antibody) is spatially coincident with the onset of IEG induction [c-fos, zif 268, and map kinase phosphatase-1 (MKP-1) detected by in situ hybridization] in the striatum, bilaterally. Both Elk-1 and CREB transcription factors (targeting SRE and CRE DNA regulatory elements, respectively) were hyperphosphorylated in register with ERK activation and IEG mRNA induction. However, their hyperphosphorylation occurred in different subcellular compartments: the cytoplasm and the nucleus for Elk-1 and the nucleus for CREB. The role of the ERK signaling cascade in gene regulation was confirmed after intrastriatal and unilateral injection of the specific ERK inhibitor PD 98059, which completely abolished c-fos, zif 268, and MKP-1 mRNA induction in the injected side. Of interest, both Elk-1 and CREB hyperphosphorylation also was impaired after PD 98059 injection. Thus two different ERK modules, one depending on the cytoplasmic activation of Elk-1 and the other one depending on the nuclear activation of CREB, control IEG transcriptional regulation in our model. Our findings provide significant insights into intracellular mechanisms underlying synaptic plasticity in the striatum.

In vivo expression and regulation of Elk-1, a target of the extracellular-regulated kinase signaling pathway, in the adult rat brain.

The transcription factor Elk-1, a nuclear target of extracellular-regulated kinases (ERKs), plays a pivotal role in immediate early gene induction by external stimuli. Notably, the degree of phosphorylation of Elk-1 is tightly correlated with the level of activation of transcription of c-fos by proliferative signals. No data yet indicate the role of Elk-1 in the adult brain in vivo. To address this question, we have analyzed in the present work (1) Elk-1 mRNA and protein expression in the adult rat brain, and (2) the regulation of Elk-1 (i.e., its phosphorylation state) in an in vivo model of immediate early gene (IEG) induction: an electrical stimulation of the cerebral cortex leading to c-fos and zif268 mRNA induction in the striatum. Using in situ hybridization, we show that Elk-1 mRNA is expressed in various brain structures of adult rat, and that this expression is exclusively neuronal. We demonstrate by immunocytochemistry using various specific Elk-1 antisera that the protein is not only nuclear (as shown previously in transiently transfected cell lines) but is also present in soma, dendrites, and axon terminals. On electrical stimulation of the glutamatergic corticostriatal pathway, we show a strict spatiotemporal correspondence among ERK activation, Elk-1 phosphorylation, and IEG mRNA induction. Furthermore, both activated proteins, analyzed by immunocytochemistry, are found in cytosolic and nuclear comparments of neuronal cells in the activated area. Our data suggest that the ERK signaling pathway plays an important role in regulating genes controlled by serum response element sites via phosphorylation of Elk-1 in vivo.

Effect of electrical stimulation of the cerebral cortex on the expression of the Fos protein in the basal ganglia.

The protein Fos is a transcription factor which is quickly induced in response to a variety of extracellular signals. Since this protein is expressed in a variety of neuronal systems in response to activation of synaptic afferents, it has been suggested that it might contribute to activity-dependent plasticity in neural networks. The present study investigated the effect of cortical electrical stimulation on the expression of Fos in the basal ganglia in the rat, a group of structures that participate in sensorimotor learning. Results show that the repetitive application of electrical shocks in restricted areas of the cerebral cortex induces an expression of Fos mostly confined to the striatum and the subthalamic nucleus. The induction which can be elicited from different cortical areas (sensorimotor, auditory and limbic areas) does not require particular temporal patterns of stimulation but rather depends on the total number of shocks delivered during a given period of time. Moreover, it appears to be rather independent of the number of spikes discharged by the activated cells. In the striatum, the distribution of immunoreactive neurons is precisely delineated and conforms to the known topographical organization of stimulated corticostriatal projections. As demonstrated using a variety of double labelling techniques (combination of the immunocytochemical detection of Fos with the autoradiography of mu opioid receptors, calbindin immunocytochemistry, in situ hybridization of preproenkephalin and preprotachykinin A messenger RNAs), striatal neurons which express Fos are mostly localized in the matrix compartment and concern equally enkephaline and substance P containing efferent neurons. In the subthalamic nucleus, Fos expression evoked by cortical stimulation is also confined to discrete regions of the nucleus, the localizations corresponding to the primary projection site of the stimulated cortical cells. These results indicate that in addition to its phasic synaptic influence on the basal ganglia, the cerebral cortex could exert a long-term effect on the functional state of this system via a genomic control. Since the basal ganglia are involved in sensorimotor learning and motor habit formation, it is tempting to speculate that the activity-dependent Fos induction at corticostriatal and subthalamic synapses may contribute to consolidate the functionality of the neuronal networks activated during the completion of given motor tasks.

Effect of angiotensin II on a spinal nociceptive reflex in the rat: receptor and mechanism of action.

The effect on thermonociceptive threshold of intrathecally (i.t.) administered angiotensin II (Ang II) was assessed in the rat tail-flick test. Rats were pretreated, 15 min earlier, with i.t. naloxone (opiate antagonist), losartan (Ang II selective antagonist at AT1 receptor) or [Sar1, Leu8] Ang II (non selective Ang II receptor antagonist) to define the mechanism of action and the nature of the receptor subtype. Ang II (0.65-6.5 nmol) induced antinociceptive effects that peaked at 1 min post-injection and returned to baseline after 5-10 min. Naloxone (10 microg) completely inhibited the response to 6.5 nmol Ang II. Losartan (65 pmol) and [Sar1, Leu8] Ang II (6.5 nmol) blocked the antinociception induced by Ang II but were inactive against [MePhe7]neurokinin B. Furthermore, losartan failed to affect the hyperalgesic responses induced by substance P (6.5 nmol) or [beta-Ala8]neurokinin A (6.5 nmol). This study provides the first functional evidence that Ang II inhibits the transmission of thermal nociceptive information through an endogenous opioid mechanism and the activation of an AT1 receptor in the rat spinal cord.