Joanne E. Nash

PDZ domains in synapse assembly and signalling

Synaptic junctions are highly specialized structures designed to promote the rapid and efficient transmission of signals from the presynaptic terminal to the postsynaptic membrane within the central nervous system. Proteins containing PDZ domains play a fundamental organizational role at both the pre- and postsynaptic plasma membranes. This review focuses on recent advances in our understanding of the mechanisms underlying the assembly of synapses in the central nervous system.

Activation of the cannabinoid receptor by Δ9-tetrahydrocannabinol reduces γ-aminobutyric acid uptake in the globus pallidus

The interaction between GABA (gamma-aminobutyric acid) and cannabinoids in the globus pallidus was investigated by evaluating the effects of delta 9-tetrahydrocannabinol on [3H]GABA uptake into slices of rat globus pallidus. delta 9-Tetrahydrocannabinol caused a concentration-dependent decrease in GABA uptake (51% decrease at 100 microM delta 9-tetrahydrocannabinol, IC50 = 18.95 microM). This effect was reversed in a concentration-dependent manner (IC50 = 11.9 microM) by the cannabinoid receptor antagonist SR 141716A (N-(piperidin-1-yl-)5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-me thyl-1 H-pyrazole-3-arboxiamidehydrochloride. SR 141716A alone did not affect GABA uptake. These results show that cannabinoid receptor activation reduces GABA uptake in the globus pallidus.

Antiparkinsonian actions of ifenprodil in the MPTP-lesioned marmoset model of Parkinson's disease.

Dopamine-replacement strategies form the basis of most symptomatic treatments for Parkinsons disease. However, since long-term dopamine-replacement therapies are characterized by many side effects, most notably dyskinesia, the concept of a nondopaminergic therapy for Parkinsons disease has attracted great interest. To date, it has proved difficult to devise a nondopaminergic therapy with efficacy comparable to that of dopamine replacement. In animal models of Parkinsons disease, loss of striatal dopamine leads to enhanced excitation of striatal NR2B-containing NMDA receptors. This is responsible, in part at least, for generating parkinsonian symptoms. Here we demonstrate that, in the MPTP-lesioned marmoset, monotherapy with the NR2B-selective NMDA receptor antagonist, ifenprodil, administered de novo, has antiparkinsonian effects equivalent to those of l-DOPA (administered as its methyl ester form). In MPTP-lesioned marmosets, median mobility scores, following vehicle-treatment were 12.5/h (range 6-21), compared to 61/h (range 26-121) in normal, non-MPTP-lesioned animals. Following ifenprodil (10 mg/kg) treatment in MPTP-lesioned marmosets, the median mobility score was 66/h (range 34-93), and following l-DOPA (10 mg/kg i.p.) treatment 89/h (range 82-92). The data support the proposal that NR2B-selective NMDA receptor antagonists have potential as a nondopaminergic monotherapy for the treatment of parkinsonian symptoms when given de novo.

The NR2B-selective NMDA receptor antagonist CP-101,606 exacerbates L-DOPA-induced dyskinesia and provides mild potentiation of anti-parkinsonian effects of L-DOPA in the MPTP-lesioned marmoset model of Parkinson's disease

In Parkinsons disease (PD), degeneration of the dopaminergic nigrostriatal pathway leads to enhanced transmission at NMDA receptors containing NR2B subunits. Previous studies have shown that some, but not all, NR2B-containing NMDA receptor antagonists alleviate parkinsonian symptoms in animal models of PD. Furthermore, enhanced NMDA receptor-mediated transmission underlies the generation of L-DOPA-induced dyskinesia (LID). The subunit content of NMDA receptors responsible for LID is not clear. Here, we assess the actions of the NMDA antagonist CP-101,606 in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned marmoset model of Parkinsons disease. CP-101,606 is selective for NMDA receptors containing NR2B subunits, with higher affinity for NR1/NR2B complexes compared to ternary NR1/NR2A/NR2B complexes. CP-101,606 had no significant effect on parkinsonian symptoms when administered as monotherapy over a range of doses (0.1-10 mg/kg). CP-101,606 provided a modest potentiation of the anti-parkinsonian actions of L-DOPA (8 mg/kg), although, at doses of 1 and 3 mg/kg, CP-101,606 exacerbated LID. Results of this study provide further evidence of differences in the anti-parkinsonian activity and effects on LID of the NR2B subunit selective NMDA receptor antagonists. These distinctions may reflect disparities in action on NR1/NR2B as opposed to NR1/NR2A/NR2B receptors.

Subcellular redistribution of the synapse-associated proteins PSD-95 and SAP97 in animal models of Parkinson’s disease and L-DOPA-induced dyskinesia

Abnormalities in subcellular localization and interaction between receptors and their signaling molecules occur within the striatum in Parkinson&s disease (PD) and L‐DOPA‐induced dyskinesia (LID). Synapse‐associated proteins (SAPs), for example, PSD‐95 and SAP97 organize the molecular architecture of synapses and regulate interactions between receptors and downstream‐signaling molecules. Here, we show that expression and subcellular distribution of PSD‐95 and SAP97 are altered in the striatum of unilateral 6‐OHDA‐lesioned rats following repeated vehicle (a model of PD) or L‐DOPA administration (a model of L‐DOPA‐induced dyskinesia). Furthermore, following dopamine‐depletion and development of behavioral deficits in Rotorod performance, indicative of parkinsonism, we observed a dramatic decrease in total striatal levels of PSD‐95 and SAP97 (to 25.6 ± 9.9% and 19.0 ± 5.0% of control, respectively). The remaining proteins were redistributed from the synapse into vesicular compartments. L‐DOPA (6.5mg/kg twice a day, 21 days) induced a rotational response, which became markedly enhanced with repeated treatment (day 1: ‒15.8±7.3 rotations cf day 21: 758.2±114.0 rotations). Post L‐DOPA treatment, PSD‐95 and SAP97 levels increased (367.4 ± 43.2% and 159.9 ± 9.5% from control values, respectively), with both being redistributed toward synaptic membranes from vesicular compartments. In situ hybridization showed that changes in total levels of PSD‐95, but not SAP97, were accompanied by qualitatively similar changes in mRNA. These data highlight the potential role of abnormalities in the subcellular distribution of SAPs in the pathophysiology of a neurological disease.

Characterisation of striatal NMDA receptors involved in the generation of parkinsonian symptoms: intrastriatal microinjection studies in the 6-OHDA-lesioned rat.

Treatments for Parkinsons disease based on replacement of lost dopamine have several problems. Following loss of dopamine, enhanced N‐methyl‐D‐aspartate (NMDA) receptor‐mediated transmission in the striatum is thought to be part of the cascade of events leading to the generation of parkinsonian symptoms. We determined the localisation and pharmacological characteristics of NMDA receptors that play a role in generating parkinsonian symptoms within the striatum. Rats were lesioned unilaterally with 6‐hydroxydopamine (6‐OHDA), and cannulae implanted bilaterally to allow injection of a range of NMDA receptor antagonists at different striatal sites. When injected rostrally into the dopamine‐depleted striatum, the glycine site partial agonist, (+)‐HA‐966 (44–400 nmol) caused a dose‐dependent contraversive rotational response consistent with an antiparkinsonian action. (+)‐HA‐966 (400 nmol) had no effect when infused into more caudal regions of the dopamine‐depleted striatum, or following injection into any striatal region on the dopamine‐intact side. To determine the pharmacological profile of NMDA receptors involved in inducing parkinsonism in 6‐OHDA‐lesioned rats, a range of NMDA receptor antagonists was infused directly into the rostral striatum. Ifenprodil (100 nmol) and 7‐chlorokynurenate (37 nmol), but not MK‐801 (15 nmol) or D‐APV (25 nmol) elicited a dramatic rotational response when injected into the dopamine‐depleted striatum. This pharmacological profile is not consistent with an effect mediated via blocking NR2B‐containing NMDA receptors. The effect of intrastriatal injection of ifenprodil was increased in animals previously treated with levodopa (L‐dopa) methyl ester. This was seen as an increase in on‐time and in peak rotational response. We propose that stimulation of NR2B‐containing NMDA receptors in the rostral striatum underlies the generation of parkinsonian symptoms. These studies are in line with previous findings suggesting that administration of NR2B‐selective NMDA receptor antagonists may be therapeutically beneficial for parkinsonian patients, when given de novo and following L‐dopa treatment.

Selective loss of bi-directional synaptic plasticity in the direct and indirect striatal output pathways accompanies generation of parkinsonism and l-DOPA induced dyskinesia in mouse models

Parkinsonian symptoms arise due to over-activity of the indirect striatal output pathway, and under-activity of the direct striatal output pathway. l-DOPA-induced dyskinesia (LID) is caused when the opposite circuitry problems are established, with the indirect pathway becoming underactive, and the direct pathway becoming over-active. Here, we define synaptic plasticity abnormalities in these pathways associated with parkinsonism, symptomatic benefits of l-DOPA, and LID. We applied spike-timing dependent plasticity protocols to cortico-striatal synapses in slices from 6-OHDA-lesioned mouse models of parkinsonism and LID, generated in BAC transgenic mice with eGFP targeting the direct or indirect output pathways, with and without l-DOPA present. In naïve mice, bidirectional synaptic plasticity, i.e. LTP and LTD, was induced, resulting in an EPSP amplitude change of approximately 50% in each direction in both striatal output pathways, as shown previously. In parkinsonism and dyskinesia, both pathways exhibited unidirectional plasticity, irrespective of stimulation paradigm. In parkinsonian animals, the indirect pathway only exhibited LTP (LTP protocol: 143.5±14.6%; LTD protocol 177.7±22.3% of baseline), whereas the direct pathway only showed LTD (LTP protocol: 74.3±4.0% and LTD protocol: 63.3±8.7%). A symptomatic dose of l-DOPA restored bidirectional plasticity on both pathways to levels comparable to naïve animals (Indirect pathway: LTP protocol: 124.4±22.0% and LTD protocol: 52.1±18.5% of baseline. Direct pathway: LTP protocol: 140.7±7.3% and LTD protocol: 58.4±6.0% of baseline). In dyskinesia, in the presence of l-DOPA, the indirect pathway exhibited only LTD (LTP protocol: 68.9±21.3% and LTD protocol 52.0±14.2% of baseline), whereas in the direct pathway, only LTP could be induced (LTP protocol: 156.6±13.2% and LTD protocol 166.7±15.8% of baseline). We conclude that normal motor control requires bidirectional plasticity of both striatal outputs, which underlies the symptomatic benefits of l-DOPA. Switching from bidirectional to unidirectional plasticity drives global changes in striatal pathway excitability, and underpins parkinsonism and dyskinesia.

Disruption of the interaction between myosin VI and SAP97 is associated with a reduction in the number of AMPARs at hippocampal synapses

J. Neurochem. (2010) 112, 677–690.

LTP in hippocampal neurons is associated with a CaMKII-mediated increase in GluA1 surface expression.

J. Neurochem. (2011) 116, 530–543.

Altered function of glutamatergic cortico-striatal synapses causes output pathway abnormalities in a chronic model of parkinsonism.

OBJECTIVE In Parkinsons disease, chronic striatal dopamine depletion results in over-activity and under-activity of the indirect and direct striatal output pathways respectively. In this study, we investigated changes in the function of glutamatergic cortico-striatal synapses that contribute to abnormalities in striatal efferents. METHODS Whole-cell recordings were performed in striatal slices prepared from adult bacterial artificial chromosome mice, chronically lesioned with 6-hydroxydopamine (6-OHDA). Paired pulse facilitation, spontaneous synaptic activity, the ratio of AMPAR to NMDAR-mediated components of excitatory postsynaptic currents, AMPAR and NMDAR kinetics, current-voltage relationship and intrinsic membrane properties were assessed in indirect and direct pathway medium spiny neurons (MSNs), which were identified on the basis of expression of GFP, driven by the promoters of A2A or D1 receptor expression. The trajectory of striatal efferents, with respect to selective targeting of the globus pallidus and substantia nigra was also compared in sham-operated versus 6-OHDA-lesioned mice. RESULTS Dopamine depletion did not affect the number of pathway specific output neurons or the trajectory of striatal outputs. In sham-operated animals, cortico-striatal synapses of both striatal efferent populations exhibited paired pulse facilitation and similar ratios of AMPAR to NMDAR-mediated components of excitatory postsynaptic currents. Following striatal dopamine depletion, indirect pathway neurons exhibited decreased levels of paired pulse facilitation, enhanced sensitivity to presynaptic stimulation and an increase in the relative contribution of NMDAR to the EPSC but no change in spontaneous synaptic activity. In sham-operated mice, neurons of the direct pathway exhibited lower firing frequency compared to the indirect pathway following current injection. However, in 6-OHDA-lesioned mice, in the direct pathway, firing threshold was reduced, spike frequency adaptation developed and the frequency of spontaneous activity was also reduced. In addition, changes in the properties of NMDAR kinetics suggest that these receptors were desensitised. DISCUSSION Increased synchronicity between pre and postsynaptic neurons, as indicated by decreased paired pulse facilitation, and increased sensitivity to extracellular stimulation, combined with an increase in the contribution of NMDARs to the EPSC at cortico-striatal synapses, may contribute to the over-activity of indirect pathway neurons in the parkinsonian striatum. In contrast, a decrease in spontaneous activity, postsynaptic desensitisation to excitatory stimuli and spike frequency adaptation of cortico-striatal synapses may underlie under-activity of the direct pathway.