SECRETARIA DE TRABAJO Resolución Nº 1060/2011

In addition to receptor level changes, LID has also been linked to changes in synaptic plasticity. Synaptic plasticity includes the three mechanisms long-term potentiation (LTP), long-term depression (LTD), and depotentiation.

LTP refers to the long-lasting potentiation of the synaptic strength between two neurons as the result of a previous short, but intense, synaptic activation and agonism of specific receptors (Bliss, 2003). LTP has been induced in cortico-striatal fibers in vitro by applying high frequency stimulation (HFS) whilst activating NMDA receptors by removing magnesium from the bathing solution the tissue is kept in (Calabresi et al, 1992, 2007; Centonze et al, 2003; Picconi et al, 2003). The success of the LTP induction can be

38 demonstrated by recording the excitatory post-synaptic potentials (EPSPs) elicited in the post-synaptic neuron following later application of HFS stimulation. The magnitude of EPSPs is greater post- than pre-LTP induction (Picconi et al, 2003).

LTD refers to a long lasting decrease in the synaptic strength between two neurons. LTD has been induced in cortico-striatal fibers in vitro by applying HFS to the tissue while NMDA receptors are inactivated by adding external magnesium to the bathing solution the tissue is kept in (Centonze et al, 2003). Like LTP, the presence of LTD may be measured by recording EPSPs or field potentials. Following LTD induction these are of lower magnitude than before induction (Bliss et al, 2003; Pisani et al, 2005; Calaresi et al, 1992a,b).

Following successful striatal LTP induction in cortico-striatal fibres, the potentiated response can be reversed in vitro by applying LFS to the presynaptic neuron. This process is known as depotentiation. Unlike LTD it does not represent a shift away from the baseline synaptic strength, but instead a reversal of the LTP induced potentiation (Picconi et al, 1992; Bliss et al, 2003; Pisani et al, 2005: Huang et al, 2010).

In vitro experiments have enabled a greater understanding of the induction of cortico-

striatal LTP and LTD. Both forms of plasticity are blocked in cortico-striatal tissue from rats that have undergone 6-OHDA lesions (Calabresi et al, 1992a; Kerr & Wickens, 2001),

suggesting a role of dopamine in both forms of plasticity. By using pharmacological blockade paradigms further insight into the receptors implicated in plasticity has been gained. Cortico- striatal LTP has been shown to depend on co-activation of mGlu1 & mGlu5 receptors (Gubellini et al, 2003), NMDA receptors (Calabresi et al, 1992b), and D1 or D5 receptors (

Table 1.2; Kerr & Wickens, 2001). Cortico-striatal LTD formation, on the other hand, has been shown to be dependent on activation of mGlu1 receptors, D5 receptors, and D2-type receptors (

Table 1.2; Calabresi et al, 1992a; Gubellini et al, 2001; Centonze et al, 2003). Cortico-striatal depotentiation has been induced in vitro in the absence of magnesium (Picconi et al, 2003) which suggests that its induction involves NMDA activation.

Furthermore, depotentiation has also been shown to require activation of protein phosphates 1 and 2A, and to be inhibited by D1 agonism (Picconi et al, 2003).

While cortico-striatal LTP and LTD are typically blocked in tissue from rats that have received 6-OHDA lesions (Calabresi et al, 1992a; Kerr & Wickens, 2001), in vitro data have demonstrated restored ability for LTP formation in tissue from lesion rats that were chronically treated with L-DOPA prior to culling (Picconi et al, 2003). Interestingly, the data demonstrated a difference in the ability for cortico-striatal plasticity in tissue from lesion rats

39 that did and did not develop LID in response to chronic L-DOPA treatment. Specifically, depotentiation of LTP could be induced in cortico-striatal tissue from L-DOPA treated non- dyskinetic rats, but not in tissue from rats that developed LID in response to the L-DOPA treatment (Picconi et al, 2003). This is believed to be due to the increased phosphorylation of

Table 1.2. An overview of the receptors that have been implicated in the induction of cortico-striatal long-term potentiation (LTP) and long-term depression (LTD).

mGlu1 mGlu5 NMDA D1 D5 D2 type

LTP x x x x x

LTD x x X

DARPP-32 that is associated with LID (e.g. Santini et al, 2007). Data obtained from sham lesion rats have shown that phosphorylation of DARPP-32, which is induced by D1 receptor activation, inhibits phosphatase 1 which is required for cortico-striatal depotentiation to occur (Picconi et al, 2003). Hence, the phosphorylation of DARP32 that is known to occur in LID may underlie the loss of cortico-striatal depotentiation observed in tissue from lesion rats that have developed LID (Picconi et al, 2003).

Clinical data suggest that the disrupted synaptic plasticity observed in preclinical models may translate to the PD patients. In the clinic, LTP like plasticity may be induced in the motor cortex using protocols involving transmagnetic stimulation (TMS). One such protocol is the paired associative stimulation (PAS) method in which TMS applied over the motor cortex is paired with median nerve stimulation at the wrist. Changes in the amplitude of motor evoked potentials (MEPs) at the abductor pollicis brevis is then used as an indicator of potentiated responses. Using this protocol, Morgante and colleagues (2006) demonstrated a loss of LTP like plasticity in Parkinsonian patients tested off-medication. This plasticity was restored by acute administration of dopaminergic medication in non-dyskinetic, but not in dyskinetic, patients (Morgante et al, 2006). In line with preclinical findings (e.g. Picconi et

al, 2003) the data thus demonstrated a requirement for dopamine for LTP like plasticity to

occur in the cortico-striatal circuitry. However, the findings reported by Morgante and colleagues (2006) differed from preclinical data (Picconi et al, 2003) by being unable to induce LTP-like plasticity in dyskinetic participants. One possible reason for the discrepancy is the protocols used: whereas preclinical experiments (e.g. Picconi et al, 2003) study LTP directly in cortico-striatal tissue, Morgante and colleagues‟ (2006) protocol only measured

40 LTP indirectly by measuring the effects of TMS on motor MEPs. There was thus a difference both in the methodology used and the specific brain structures being studied.

A later study readdressed the issue using a new protocol that had been demonstrated to be more consistent in its ability to induce LTP like effects in PD patients (Huang et al, 2010, 2011). With this protocol, it was also possible to study both LTP like and depotentiation like plasticity (Huang et al, 2010, Huang et al, 2011). In their study, Huang and colleagues (2011) were able to induce LTP like potentiation in both dyskinetic and non- dyskinetic patients when these received their full L-DOPA dose. Furthermore, in line with preclinical data (Picconi et al, 2003) only the non-dyskinetic group was found to have retained the ability for depotentiation. The data from Huang and colleagues (2011) therefore suggested that preclinical findings of disrupted synaptic plasticity in a model of PD (Picconi

et al, 2003) may translate to the clinic. The data further suggested that the failure of an earlier

study (Morgante et al, 2006) to demonstrate LTP like plasticity in dyskinetic PD patients may have been due to the methodology used.

The hypothesis that subtle differences in methodology and patients medication have a large impact on the ability to induce LTP like plasticity using PAS protocol is supported by data from other clinical experiments. In the previously described study, Huang and colleagues (2011) were able to induce LTP like plasticity in PD patients receiving their full L-DOPA dose. However, they failed to induce LTP like plasticity when patients received half their dose (Huang et al, 2011). This is consistent with data from other studies where LTP-like plasticity could not be induced in patients receiving a low L-DOPA dose prior to testing (Suppa et al, 2011). Together, these studies show that the ability for inducing LTP like potentiation in patients is highly sensitive to not only the protocol (Morgante et al, 2006, Hunag et al, 2010, 2011) but also the dose of medication the participants receive (Suppa et al, 2011, Huang et al, 2011).

A weakness of the clinical studies described above (Morgante et al, 2006; Huang et

al, 2010, 2011; Suppa et al, 2011) is that they only measured LTP and depotentiation

indirectly - unlike in vitro studies which are able to directly measure the effect of potentiation on post-synaptic responses. One instance in which synaptic plasticity can be measured directly in the clinic is during deep-brain stimulation (DBS) implantation, a surgical therapy aimed to relieve PD symptoms by implantation of electrodes to the basal ganglia (Starr et al, 1998). During such implantation, Prescott and colleagues (2009) were able to record field evoked potentials (fEP) following HFS in the SNr of patients that were either on or off L-DOPA medication at the time of surgery. Prescott and colleagues (2009) demonstrated that

41 LTP could only be induced in patients who were on L-DOPA at the time of testing. Whilst recording from the SNr (Prescott et al, 2009) rather than cortico-striatal tissue as in the previously described preclinical study (Picconi et al, 2003), the data offer further support for findings of a disruptive effect of PD on basal ganglia plasticity, and the need for dopamine for successful LTP induction in the basal ganglia network, as well as suggesting that the abnormal plasticity associated with PD may affect multiple parts of the basal ganglia circuitry.

Whilst further work is required to understand the precise effect of PD and acute and chronic L-DOPA on basal ganglia synaptic plasticity, the cited studies suggest a long-term effect on synaptic plasticity on basal ganglia plasticity. These changes, together with the receptor changes also associated with L-DOPA and LID onset, could both be hypothesised to affect non-motor behaviour mediated by the striatum. In addition to motor function, the striatum is known to mediate non-motor functions such as motor learning, and goal-directed, and habitual behaviour (e.g. Wächter et al, 2010; Yin et al, 2004, 2005; Reynolds et al, 2001; Luft et al, 2004; Okulski et al, 2002). Therefore, further to the already known effect of chronic L-DOPA on the development of LID and motor fluctuations (Marsden & Parkes, 1977), chronic exposure to the drug may impact on non-motor functions.