Paola Bonsi

Impaired dopamine release and synaptic plasticity in the striatum of PINK1-deficient mice

Parkinsons disease (PD) is characterized by the selective vulnerability of the nigrostriatal dopaminergic circuit. Recently, loss-of-function mutations in the PTEN-induced kinase 1 (PINK1) gene have been linked to early-onset PD. How PINK1 deficiency causes dopaminergic dysfunction and degeneration in PD patients is unknown. Here, we investigate the physiological role of PINK1 in the nigrostriatal dopaminergic circuit through the generation and multidisciplinary analysis of PINK1−/− mutant mice. We found that numbers of dopaminergic neurons and levels of striatal dopamine (DA) and DA receptors are unchanged in PINK1−/− mice. Amperometric recordings, however, revealed decreases in evoked DA release in striatal slices and reductions in the quantal size and release frequency of catecholamine in dissociated chromaffin cells. Intracellular recordings of striatal medium spiny neurons, the major dopaminergic target, showed specific impairments of corticostriatal long-term potentiation and long-term depression in PINK1−/− mice. Consistent with a decrease in evoked DA release, these striatal plasticity impairments could be rescued by either DA receptor agonists or agents that increase DA release, such as amphetamine or l-dopa. These results reveal a critical role for PINK1 in DA release and striatal synaptic plasticity in the nigrostriatal circuit and suggest that altered dopaminergic physiology may be a pathogenic precursor to nigrostriatal degeneration.

Endogenous Serotonin Excites Striatal Cholinergic Interneurons via the Activation of 5-HT 2C, 5-HT6, and 5-HT7 Serotonin Receptors: Implications for Extrapyramidal Side Effects of Serotonin Reuptake Inhibitors

The striatum is richly innervated by serotonergic afferents from the raphe nucleus. We explored the effects of this input on striatal cholinergic interneurons from rat brain slices, by means of both conventional intracellular and whole-cell patch-clamp recordings. Bath-applied serotonin (5-HT, 3–300 μM), induced a dose-dependent membrane depolarization and increased the rate of spiking. This effect was mimicked by the 5-HT reuptake blockers citalopram and fluvoxamine. In voltage-clamped neurons, 5-HT induced an inward current, whose reversal potential was close to the K+ equilibrium potential. Accordingly, the involvement of K+ channels was confirmed either by increasing extracellular K+ concentration and by blockade of K+ channels with barium. Single-cell reverse transcriptase-polymerase chain reaction (RT-PCR) profiling demonstrated the presence of 5-HT2C, 5-HT6, and 5-HT7 receptor mRNAs in identified cholinergic interneurons. The depolarization/inward current induced by 5-HT was partially mimicked by the 5-HT2 receptor agonist 2,5-dimethoxy-4-iodoamphetamine and antagonized by both ketanserin and the selective 5-HT2C antagonist RS102221, whereas the selective 5-HT3 and 5-HT4 receptor antagonists tropisetron and RS23597-190 had no effect. The depolarizing response to 5-HT was also reduced by the selective 5-HT6 and 5-HT7 receptor antagonists SB258585 and SB269970, respectively, and mimicked by the 5-HT7 agonist, 5-CT. Accordingly, activation of either 5-HT6 or 5-HT7 receptor induced an inward current. The 5-HT response was attenuated by U73122, blocker of phospholipase C, and by SQ22,536, an inhibitor of adenylyl cyclase. These results suggest that 5-HT released by serotonergic fibers originating in the raphe nuclei has a potent excitatory effect on striatal cholinergic interneurons.

Selective involvement of mGlu1 receptors in corticostriatal LTD.

Although metabotropic glutamate receptors (mGluRs) have been proposed to play a role in corticostriatal long-term depression (LTD), the specific receptor subtype required for this form of synaptic plasticity has not been characterized yet. Thus, we utilized a corticostriatal brain slice preparation and intracellular recordings from striatal spiny neurons to address this issue. We observed that both AIDA (100 microM) and LY 367385 (30 microM), two blockers of mGluR1s, were able to fully prevent the induction of this form of synaptic plasticity, whereas MPEP (30 microM), a selective antagonist of the mGluR5 subtype, did not significantly affect the amplitude and time-course of corticostriatal LTD. Both AIDA and LY 367385 were ineffective on LTD when applied after its induction. The critical role of mGluR1s in the formation of corticostriatal LTD was confirmed in experiments performed on mice lacking mGluR1s. In these mice, in fact, a significant reduction of the LTD amplitude was observed in comparison to the normal LTD measured in their wild-type counterparts. We found that neither acute pharmacological blockade of mGluR1s nor the genetic disruption of these receptors affected the presynaptic modulation of corticostriatal excitatory postsynapic potentials (EPSPs) exerted by DCG-IV and L-SOP, selective agonists of group II and III mGluRs, respectively. Our data show that the induction of corticostriatal LTD requires the activation of mGluR1 but not mGluR5. mGluR1-mediated control of this form of synaptic plasticity may play a role in the modulatory effect exerted by mGluRs in the basal ganglia-related motor activity.

Intracellular Calcium Increase in Epileptiform Activity : Modulation by Levetiracetam and Lamotrigine

Summary:  Purpose: Alterations in neuronal calcium (Ca2+) homeostasis are believed to play an essential role in the generation and propagation of epileptiform events. Levetiracetam (LEV) and lamotrigine (LTG), novel antiepileptic drugs (AEDs), were tested on epileptiform events and the corresponding elevations in intracellular Ca2+ concentration ([Ca2+]i) recorded from rat neocortical slices.

Targeting striatal cholinergic interneurons in Parkinson’s disease: Focus on metabotropic glutamate receptors

In the early sixties, anticholinergic drugs were introduced in the pharmacological treatment of Parkinsons disease (PD). The rationale behind their utilisation in the treatment of the disease was based on the evidence of an imbalance between the dopaminergic inputs and the intrinsic cholinergic innervation within the striatum. Metabotropic glutamate (mGlu) receptors have been shown to play a key role in striatal function both in physiological conditions and in experimental models of diseases affecting this brain area. Indeed, compelling electrophysiological and morphological evidence shows that mGlu receptors are highly expressed at cellular level and exert a profound modulatory role on cholinergic interneurons excitability. This review will provide a brief survey of studies on the localization and function of mGlu receptors in cholinergic interneurons. The potential relevance of these findings in the control of motor function and in the treatment of PD will be discussed.

Plastic and behavioral abnormalities in experimental Huntington's disease: A crucial role for cholinergic interneurons

Huntingtons disease (HD) is a fatal hereditary neurodegenerative disease causing degeneration of striatal spiny neurons, whereas cholinergic interneurons are spared. This cell-type specific pathology produces an array of abnormalities including involuntary movements, cognitive impairments, and psychiatric disorders. Although the genetic mutation responsible for HD has been identified, little is known about the early synaptic changes occurring within the striatal circuitry at the onset of clinical symptoms. We therefore studied the synaptic plasticity of spiny neurons and cholinergic interneurons in two animal models of early HD. As a pathogenetic model, we used the chronic subcutaneous infusion of the mitochondrial toxin 3-nitropropionic acid (3-NP) in rats. This treatment caused striatal damage and impaired response flexibility in the cross-maze task as well as defective extinction of conditioned fear suggesting a perseverative behavior. In these animals, we observed a loss of depotentiation in striatal spiny neurons and a lack of long-term potentiation (LTP) in cholinergic interneurons. These abnormalities of striatal synaptic plasticity were also observed in R6/2 transgenic mice, a genetic model of HD, indicating that both genetic and phenotypic models of HD show cell-type specific alterations of LTP. We also found that in control rats, as well as in wild-type (WT) mice, depotentiation of spiny neurons was blocked by either scopolamine or hemicholinium, indicating that reversal of LTP requires activation of muscarinic receptors by endogenous acetylcholine. Our findings suggest that the defective plasticity of cholinergic interneurons could be the primary event mediating abnormal functioning of striatal circuits, and the loss of behavioral flexibility typical of early HD might largely depend on cell-type specific plastic abnormalities.

Loss of Muscarinic Autoreceptor Function Impairs Long-Term Depression But Not Long-Term Potentiation in the Striatum

Muscarinic autoreceptors regulate cholinergic tone in the striatum. We investigated the functional consequences of genetic deletion of striatal muscarinic autoreceptors by means of electrophysiological recordings from either medium spiny neurons (MSNs) or cholinergic interneurons (ChIs) in slices from single M4 or double M2/M4 muscarinic acetylcholine receptor (mAChR) knock-out (−/−) mice. In control ChIs, the muscarinic agonist oxotremorine (300 nm) produced a self-inhibitory outward current that was mostly reduced in M4−/− and abolished in M2/M4−/− mice, suggesting an involvement of both M2 and M4 autoreceptors. In MSNs from both M4−/− and M2/M4−/− mice, muscarine caused a membrane depolarization that was prevented by the M1 receptor-preferring antagonist pirenzepine (100 nm), suggesting that M1 receptor function was unaltered. Acetylcholine has been involved in striatal long-term potentiation (LTP) or long-term depression (LTD) induction. Loss of muscarinic autoreceptor function is predicted to affect synaptic plasticity by modifying striatal cholinergic tone. Indeed, high-frequency stimulation of glutamatergic afferents failed to induce LTD in MSNs from both M4−/− and M2/M4−/− mice, as well as in wild-type mice pretreated with the M2/M4 antagonist AF-DX384 (11-[[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5,1 1-dihydro-6H-pyrido[2,3b][1,4] benzodiazepin-6-one). Interestingly, LTD could be restored by either pirenzepine (100 nm) or hemicholinium-3 (10 μm), a depletor of endogenous ACh. Conversely, LTP induction did not show any difference among the three mouse strains and was prevented by pirenzepine. These results demonstrate that M2/M4 muscarinic autoreceptors regulate ACh release from striatal ChIs. As a consequence, endogenous ACh drives the polarity of bidirectional synaptic plasticity.

NR2B Subunit Exerts a Critical Role in Postischemic Synaptic Plasticity

Background and Purpose— We characterized the differential effect of the NR2B subunit antagonist ifenprodil in the induction of activity-dependent long-term potentiation (LTP) and of postischemic LTP as well as in the neuronal damage induced by focal ischemia. Methods— Intracellular recordings were obtained from rat corticostriatal slice preparations. High-frequency stimulation of corticostriatal fibers was used as a LTP-inducing protocol. In vitro ischemia was induced by oxygen and glucose deprivation. In vivo ischemia was induced by permanent middle cerebral artery occlusion. Intracellular recordings were also performed in the ischemic penumbra. Results— Antagonists selectively targeting N-methyl-d-aspartate receptors containing the NR2B subunit blocked postischemic LTP without affecting activity-dependent LTP. In a model of focal ischemia, blockade of NR2B subunit in vivo caused reduction of brain damage, amelioration of neurological outcome, and normalization of the synaptic levels of NR2B subunits. Moreover, the antagonism of NR2B subunit was able to rescue the activity-dependent LTP in the ischemic penumbra. Conclusions— We suggest that NR2B subunits contribute to the striatal damage caused by in vivo and in vitro ischemia and play a critical role in the induction of postischemic LTP as well as in the suppression of activity-dependent LTP in the ischemic penumbra.

Centrality of striatal cholinergic transmission in Basal Ganglia function

Work over the past two decades revealed a previously unexpected role for striatal cholinergic interneurons in the context of basal ganglia function. The recognition that these interneurons are essential in synaptic plasticity and motor learning represents a significant step ahead in deciphering how the striatum processes cortical inputs, and why pathological circumstances cause motor dysfunction. Loss of the reciprocal modulation between dopaminergic inputs and the intrinsic cholinergic innervation within the striatum appears to be the trigger for pathophysiological changes occurring in basal ganglia disorders. Accordingly, there is now compelling evidence showing profound changes in cholinergic markers in these disorders, in particular Parkinsons disease and dystonia. Based on converging experimental and clinical evidence, we provide an overview of the role of striatal cholinergic transmission in physiological and pathological conditions, in the context of the pathogenesis of movement disorders.

Impaired striatal D2 receptor function leads to enhanced GABA transmission in a mouse model of DYT1 dystonia.

DYT1 dystonia is caused by a deletion in a glutamic acid residue in the C-terminus of the protein torsinA, whose function is still largely unknown. Alterations in GABAergic signaling have been involved in the pathogenesis of dystonia. We recorded GABA- and glutamate-mediated synaptic currents from a striatal slice preparation obtained from a mouse model of DYT1 dystonia. In medium spiny neurons (MSNs) from mice expressing human mutant torsinA (hMT), we observed a significantly higher frequency, but not amplitude, of GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) and miniature currents (mIPSCs), whereas glutamate-dependent spontaneous excitatory synaptic currents (sEPSCs) were normal. No alterations were found in mice overexpressing normal human torsinA (hWT). To identify the possible sources of the increased GABAergic tone, we recorded GABAergic Fast-Spiking (FS) interneurons that exert a feed-forward inhibition on MSNs. However, both sEPSC and sIPSC recorded from hMT FS interneurons were comparable to hWT and non-transgenic (NT) mice. In physiological conditions, dopamine (DA) D2 receptor act presynaptically to reduce striatal GABA release. Of note, application of the D2-like receptor agonist quinpirole failed to reduce the frequency of sIPSCs in MSNs from hMT as compared to hWT and NT mice. Likewise, the inhibitory effect of quinpirole was lost on evoked IPSCs both in MSNs and FS interneurons from hMT mice. Our findings demonstrate a disinhibition of striatal GABAergic synaptic activity, that can be at least partially attributed to a D2 DA receptor dysfunction.