Disease mechanisms as Subtypes: Mitochondrial and bioenergetic dysfunction.

Parkinson disease (PD) is the second most common neurodegenerative disease in the world. Despite its enormous human and societal cost, there is no disease-modifying therapy for PD. This unmet medical need reflects our limited understanding of PD pathogenesis. One of the most important clues comes from the recognition that PD motor symptoms arises from the dysfunction and degeneration of a very select group of neurons in the brain. These neurons have a distinctive set of anatomic and physiologic traits that reflect their role in brain function. These traits elevate mitochondrial stress, potentially making them particularly vulnerable to age, as well as to genetic mutations and environmental toxins linked to PD incidence. In this chapter, the literature supporting this model is outlined, along with gaps in our knowledge base. The translational implications of this hypothesis are then discussed, with a focus on why disease-modification trials have failed to date and what this means for the development of new strategies for altering disease course.

Ca2+ channels couple spiking to mitochondrial metabolism in substantia nigra dopaminergic neurons.

How do neurons match generation of adenosine triphosphate by mitochondria to the bioenergetic demands of regenerative activity? Although the subject of speculation, this coupling is still poorly understood, particularly in neurons that are tonically active. To help fill this gap, pacemaking substantia nigra dopaminergic neurons were studied using a combination of optical, electrophysiological, and molecular approaches. In these neurons, spike-activated calcium (Ca2+) entry through Cav1 channels triggered Ca2+ release from the endoplasmic reticulum, which stimulated mitochondrial oxidative phosphorylation through two complementary Ca2+-dependent mechanisms: one mediated by the mitochondrial uniporter and another by the malate-aspartate shuttle. Disrupting either mechanism impaired the ability of dopaminergic neurons to sustain spike activity. While this feedforward control helps dopaminergic neurons meet the bioenergetic demands associated with sustained spiking, it is also responsible for their elevated oxidant stress and possibly to their decline with aging and disease.

Cholinergic Interneurons Amplify Thalamostriatal Excitation of Striatal Indirect Pathway Neurons in Parkinson's Disease Models.

The motor symptoms of Parkinson's disease (PD) are thought to stem from an imbalance in the activity of striatal direct- and indirect-pathway spiny projection neurons (SPNs). Disease-induced alterations in the activity of networks controlling SPNs could contribute to this imbalance. One of these networks is anchored by the parafascicular nucleus (PFn) of the thalamus. To determine the role of the PFn in striatal PD pathophysiology, optogenetic, chemogenetic, and electrophysiological tools were used in ex vivo slices from transgenic mice with region-specific Cre recombinase expression. These studies revealed that in parkinsonian mice, the functional connectivity of PFn neurons with indirect pathway SPNs (iSPNs) was selectively enhanced by cholinergic interneurons acting through presynaptic nicotinic acetylcholine receptors (nAChRs) on PFn terminals. Attenuating this network adaptation by chemogenetic or genetic strategies alleviated motor-learning deficits in parkinsonian mice, pointing to a potential new therapeutic strategy for PD patients.

Heterosynaptic regulation of external globus pallidus inputs to the subthalamic nucleus by the motor cortex.

The two principal movement-suppressing pathways of the basal ganglia, the so-called hyperdirect and indirect pathways, interact within the subthalamic nucleus (STN). An appropriate level and pattern of hyperdirect pathway cortical excitation and indirect pathway external globus pallidus (GPe) inhibition of the STN are critical for normal movement and are greatly perturbed in Parkinson's disease. Here we demonstrate that motor cortical inputs to the STN heterosynaptically regulate, through activation of postsynaptic NMDA receptors, the number of functional GABAA receptor-mediated GPe-STN inputs. Therefore, a homeostatic mechanism, intrinsic to the STN, balances cortical excitation by adjusting the strength of GPe inhibition. However, following the loss of dopamine, excessive cortical activation of STN NMDA receptors triggers GPe-STN inputs to strengthen abnormally, contributing to the emergence of pathological, correlated activity.

CaV1.3-selective L-type calcium channel antagonists as potential new therapeutics for Parkinson's disease.

L-type calcium channels expressed in the brain are heterogeneous. The predominant class of L-type calcium channels has a Ca(V)1.2 pore-forming subunit. L-type calcium channels with a Ca(V)1.3 pore-forming subunit are much less abundant, but have been implicated in the generation of mitochondrial oxidant stress underlying pathogenesis in Parkinson's disease. Thus, selectively antagonizing Ca(V)1.3 L-type calcium channels could provide a means of diminishing cell loss in Parkinson's disease without producing side effects accompanying general antagonism of L-type calcium channels. However, there are no known selective antagonists of Ca(V)1.3 L-type calcium channel. Here we report high-throughput screening of commercial and 'in-house' chemical libraries and modification of promising hits. Pyrimidine-2,4,6-triones were identified as a potential scaffold; structure-activity relationship-based modification of this scaffold led to 1-(3-chlorophenethyl)-3-cyclopentylpyrimidine-2,4,6-(1H,3H,5H)-trione (8), a potent and highly selective Ca(V)1.3 L-type calcium channel antagonist. The biological relevance was confirmed by whole-cell patch-clamp electrophysiology. These studies describe the first highly selective Ca(V)1.3 L-type calcium channel antagonist and point to a novel therapeutic strategy for Parkinson's disease.

Thalamic contributions to Basal Ganglia-related behavioral switching and reinforcement.

Although the existence of prominent connections between the intralaminar thalamic nuclei and the basal ganglia has long been established, the limited knowledge of the functional relevance of this network has considerably hampered progress in our understanding of the neural mechanisms by which the thalamostriatal system integrates and regulates the basal ganglia circuitry. In this brief commentary, we will address this gap of knowledge through a discussion of the key points of a symposium entitled "Thalamic Contributions to Basal Ganglia-Related Behavioral Switching and Reinforcement" that will be presented at the 2011 Society for Neuroscience meeting. Recent anatomical and physiological data that support the role of the thalamostriatal system in action selection, attentional shifting, and reinforcement will be discussed. We will also address the possibility that degeneration of the thalamostriatal system could underlie some of the deficits in redirection of attention in response to salient stimuli seen in Parkinson's disease.

Corticospinal-specific HCN expression in mouse motor cortex: I(h)-dependent synaptic integration as a candidate microcircuit mechanism involved in motor control.

Motor cortex is a key brain center involved in motor control in rodents and other mammals, but specific intracortical mechanisms at the microcircuit level are largely unknown. Neuronal expression of hyperpolarization-activated current (I(h)) is cell class specific throughout the nervous system, but in neocortex, where pyramidal neurons are classified in various ways, a systematic pattern of expression has not been identified. We tested whether I(h) is differentially expressed among projection classes of pyramidal neurons in mouse motor cortex. I(h) expression was high in corticospinal neurons and low in corticostriatal and corticocortical neurons, a pattern mirrored by mRNA levels for HCN1 and Trip8b subunits. Optical mapping experiments showed that I(h) attenuated glutamatergic responses evoked across the apical and basal dendritic arbors of corticospinal but not corticostriatal neurons. Due to I(h), corticospinal neurons resonated, with a broad peak at ∼4 Hz, and were selectively modulated by α-adrenergic stimulation. I(h) reduced the summation of short trains of artificial excitatory postsynaptic potentials (EPSPs) injected at the soma, and similar effects were observed for short trains of actual EPSPs evoked from layer 2/3 neurons. I(h) narrowed the coincidence detection window for EPSPs arriving from separate layer 2/3 inputs, indicating that the dampening effect of I(h) extended to spatially disperse inputs. To test the role of corticospinal I(h) in transforming EPSPs into action potentials, we transfected layer 2/3 pyramidal neurons with channelrhodopsin-2 and used rapid photostimulation across multiple sites to synaptically drive spiking activity in postsynaptic neurons. Blocking I(h) increased layer 2/3-driven spiking in corticospinal but not corticostriatal neurons. Our results imply that I(h)-dependent synaptic integration in corticospinal neurons constitutes an intracortical control mechanism, regulating the efficacy with which local activity in motor cortex is transferred to downstream circuits in the spinal cord. We speculate that modulation of I(h) in corticospinal neurons could provide a microcircuit-level mechanism involved in translating action planning into action execution.

Selective participation of somatodendritic HCN channels in inhibitory but not excitatory synaptic integration in neurons of the subthalamic nucleus.

The activity patterns of subthalamic nucleus (STN) neurons are intimately linked to motor function and dysfunction and arise through the complex interaction of intrinsic properties and inhibitory and excitatory synaptic inputs. In many neurons, hyperpolarization-activated cyclic nucleotide-gated (HCN) channels play key roles in intrinsic excitability and synaptic integration both under normal conditions and in disease states. However, in STN neurons, which strongly express HCN channels, their roles remain relatively obscure. To address this deficit, complementary molecular and cellular electrophysiological, imaging, and computational approaches were applied to the rat STN. Molecular profiling demonstrated that individual STN neurons express mRNA encoding several HCN subunits, with HCN2 and 3 being the most abundant. Light and electron microscopic analysis showed that HCN2 subunits are strongly expressed and distributed throughout the somatodendritic plasma membrane. Voltage-, current-, and dynamic-clamp analysis, two-photon Ca(2+) imaging, and computational modeling revealed that HCN channels are activated by GABA(A) receptor-mediated inputs and thus limit synaptic hyperpolarization and deinactivation of low-voltage-activated Ca(2+) channels. Although HCN channels also limited the temporal summation of EPSPs, generated through two-photon uncaging of glutamate, this action was largely shunted by GABAergic inhibition that was necessary for HCN channel activation. Together the data demonstrate that HCN channels in STN neurons selectively counteract GABA(A) receptor-mediated inhibition arising from the globus pallidus and thus promote single-spike activity rather than rebound burst firing.

Thalamic gating of corticostriatal signaling by cholinergic interneurons.

Salient stimuli redirect attention and suppress ongoing motor activity. This attentional shift is thought to rely upon thalamic signals to the striatum to shift cortically driven action selection, but the network mechanisms underlying this interaction are unclear. Using a brain slice preparation that preserved cortico- and thalamostriatal connectivity, it was found that activation of thalamostriatal axons in a way that mimicked the response to salient stimuli induced a burst of spikes in striatal cholinergic interneurons that was followed by a pause lasting more than half a second. This patterned interneuron activity triggered a transient, presynaptic suppression of cortical input to both major classes of principal medium spiny neuron (MSN) that gave way to a prolonged enhancement of postsynaptic responsiveness in striatopallidal MSNs controlling motor suppression. This differential regulation of the corticostriatal circuitry provides a neural substrate for attentional shifts and cessation of ongoing motor activity with the appearance of salient environmental stimuli.

Antagonizing L-type Ca2+ channel reduces development of abnormal involuntary movement in the rat model of L-3,4-dihydroxyphenylalanine-induced dyskinesia.

BACKGROUND: Chronic L-3,4-dihydroxyphenylalanine (L-DOPA) treatment of Parkinson's disease (PD) leads to debilitating involuntary movements, termed L-DOPA-induced dyskinesia. Striatofugal medium spiny neurons (MSN) lose their dendritic spines and cortico-striatal glutamatergic synapses in PD and in experimental models of DA depletion. This loss of connectivity is triggered by a dysregulation of intraspine Cav1.3 L-type Ca2+ channels. Here we address the possible implication of DA denervation-induced spine pruning in the development of L-DOPA-induced dyskinesia. METHODS: The L-type Ca2+ antagonist, isradipine was subcutaneously delivered to rats at the doses of .05, .1, or .2 mg/kg/day, for 4 weeks, starting the day after a unilateral nigrostriatal 6-hydroxydopamine (6-OHDA) lesion. Fourteen days later, L-DOPA treatment was initiated. RESULTS: Isradipine-treated animals displayed a dose-dependent reduction in L-DOPA-induced rotational behavior and abnormal involuntary movements. Dendritic spine counting at electron microscopy level showed that isradipine (.2 mg/kg/day) prevented the 6-OHDA-induced spine loss and normalized preproenkephalin-A messenger RNA expression. Involuntary movements were not reduced when isradipine treatment was started concomitantly with L-DOPA. CONCLUSIONS: These results indicate that isradipine, at a therapeutically relevant dose, might represent a treatment option for preventing L-DOPA-induced dyskinesia in PD.

Dichotomous dopaminergic control of striatal synaptic plasticity.

At synapses between cortical pyramidal neurons and principal striatal medium spiny neurons (MSNs), postsynaptic D1 and D2 dopamine (DA) receptors are postulated to be necessary for the induction of long-term potentiation and depression, respectively-forms of plasticity thought to underlie associative learning. Because these receptors are restricted to two distinct MSN populations, this postulate demands that synaptic plasticity be unidirectional in each cell type. Using brain slices from DA receptor transgenic mice, we show that this is not the case. Rather, DA plays complementary roles in these two types of MSN to ensure that synaptic plasticity is bidirectional and Hebbian. In models of Parkinson's disease, this system is thrown out of balance, leading to unidirectional changes in plasticity that could underlie network pathology and symptoms.

Corticostriatal and thalamostriatal synapses have distinctive properties.

The two principal excitatory glutamatergic inputs to striatal medium spiny neurons (MSNs) arise from neurons in the cerebral cortex and thalamus. Although there have been many electrophysiological studies of MSN glutamatergic synapses, little is known about how corticostriatal and thalamostriatal synapses differ. Using mouse brain slices that allowed each type of synapse to be selectively activated, electrophysiological approaches were used to characterize their properties in identified striatopallidal and striatonigral MSNs. At corticostriatal synapses, a single afferent volley increased the glutamate released by a subsequent volley, leading to enhanced postsynaptic depolarization with repetitive stimulation. This was true for both striatonigral and striatopallidal MSNs. In contrast, at thalamostriatal synapses, a single afferent volley decreased glutamate released by a subsequent volley, leading to a depressed postsynaptic depolarization with repetitive stimulation. Again, this response pattern was the same in striatonigral and striatopallidal MSNs. These differences in release probability and short-term synaptic plasticity suggest that corticostriatal and thalamostriatal projection systems code information in temporally distinct ways, constraining how they regulate striatal circuitry.

Dendritic excitability of mouse frontal cortex pyramidal neurons is shaped by the interaction among HCN, Kir2, and Kleak channels.

Dendritically placed, voltage-sensitive ion channels are key regulators of neuronal synaptic integration. In several cell types, hyperpolarization/cyclic nucleotide gated (HCN) cation channels figure prominently in dendritic mechanisms controlling the temporal summation of excitatory synaptic events. In prefrontal cortex, the sustained activity of pyramidal neurons in working memory tasks is thought to depend on the temporal summation of dendritic excitatory inputs. Yet we know little about how this is accomplished in these neurons and whether HCN channels play a role. To gain a better understanding of this process, layer V-VI pyramidal neurons in slices of mouse prelimbic and infralimbic cortex were studied. Somatic voltage-clamp experiments revealed the presence of rapidly activating and deactivating cationic currents attributable to HCN1/HCN2 channels. These channels were open at the resting membrane potential and had an apparent half-activation voltage near -90 mV. In the same voltage range, K+ currents attributable to Kir2.2/2.3 and K+-selective leak (Kleak) channels were prominent. Computer simulations grounded in the biophysical measurements suggested a dynamic interaction among Kir2, Kleak, and HCN channel currents in shaping membrane potential and the temporal integration of synaptic potentials. This inference was corroborated by experiment. Blockade of Kir2/Kleak channels caused neurons to depolarize, leading to the deactivation of HCN channels, the initiation of regular spiking (4-5 Hz), and enhanced temporal summation of EPSPs. These studies show that HCN channels are key regulators of synaptic integration in prefrontal pyramidal neurons but that their functional contribution is dependent on a partnership with Kir2 and Kleak channels.