Stephen Sammut

Inhibition of Phosphodiesterase 10A Increases the Responsiveness of Striatal Projection Neurons to Cortical Stimulation

The cyclic nucleotide phosphodiesterase 10A (PDE10A) is highly expressed in striatal medium-sized spiny projection neurons (MSNs), apparently playing a critical role in the regulation of both cGMP and cAMP signaling cascades. Genetic disruption or pharmacological inhibition of PDE10A reverses behavioral abnormalities associated with subcortical hyperdopaminergia. Here, we investigate the effect of PDE10A inhibition on the activity of MSNs using single-unit extracellular recordings performed in the dorsal striatum of anesthetized rats. Antidromic stimulation of the substantia nigra pars reticulata was used to identify striatonigral (SNr+) MSNs. Intrastriatal infusion of the selective PDE10A inhibitors papaverine or TP-10 [2-{4-[-pyridin-4-yl-1-(2,2,2-trifluoroethyl)-1H-pyrazol-3-yl]-phenoxymethyl}-quinoline succinic acid] by reverse microdialysis did not affect spontaneous firing but robustly increased measures of cortically evoked spike activity in a stimulus intensity-dependent manner. Systemic administration of TP-10 also increased cortically evoked spike activity in a stimulus intensity- and dose-dependent manner. A robust increase in cortically evoked activity was apparent in SNr- MSNs (primarily striatopallidal). It is interesting that TP-10 administration did not affect cortically evoked activity in SNr+ MSNs. However, TP-10 administration increased the incidence of antidromically activated (i.e., SNr+) MSNs. These findings indicate that inhibition of striatal PDE10A activity increases the responsiveness of MSNs to depolarizing stimuli. Furthermore, given the lack of effect of TP-10 on SNr+ MSNs, we speculate that PDE10A inhibition may have a greater facilitatory effect on corticostriatal synaptic activity in striatopallidal MSNs. These data support further investigation of selective targeting of PDE signaling pathways in MSN subpopulations because this may represent a promising novel approach for treating brain disorders involving dysfunctional glutamatergic and dopaminergic neurotransmission.

Phasic Dopaminergic Transmission Increases NO Efflux in the Rat Dorsal Striatum via a Neuronal NOS and a Dopamine D 1/5 Receptor-Dependent Mechanism

Dysfunctional neurotransmission within striatal networks is believed to underlie the pathophysiology of several neurological and psychiatric disorders. Nitric oxide (NO)-producing interneurons have been shown to play a critical role in modulating striatal synaptic transmission. These interneurons receive synaptic contacts from midbrain dopamine (DA) neurons and may be regulated by DA receptor activation. In the current study, striatal NO efflux was measured in anesthetized male rats using an NO-selective electrochemical microsensor and the role of DA in modulating NO synthase (NOS) activity was assessed during electrical or chemical (bicuculline) stimulation of the substantia nigra (SN). Electrical stimuli were patterned to approximate the natural single spike or burst firing activity of midbrain DA neurons. Electrical stimulation of the SN at low frequencies induced modest increases in striatal NO efflux. In contrast, train stimulation of the SN robustly increased NO efflux in a stimulus intensity-dependent manner. NO efflux evoked by SN stimulation was similar in chloral hydrate- and urethane-anesthetized rats. The facilitatory effect of train stimulation on striatal NO efflux was transient and attenuated by systemic administration of the neuronal NOS inhibitor 7-nitroindazole and the nonselective NOS inhibitor methylene blue. Moreover, the increase in NO efflux observed during chemical and train stimulation of the SN was attenuated following systemic administration of the DA D1/5 receptor antagonist SCH 23390. SCH 23390 also blocked NO efflux induced by systemic administration of the D1/5 agonist SKF 81297. These results indicate that neuronal NOS is activated in vivo by nigrostriatal DA cell burst firing via a DA D1/5-like receptor-dependent mechanism.

Dopamine D2 receptor-dependent modulation of striatal NO synthase activity

RationaleStriatal nitric oxide (NO)-producing interneurons receive synaptic contacts from midbrain dopamine (DA) neurons and are regulated by phasic DA transmission. Classic antipsychotic drugs elevate neuronal NO synthase (NOS) expression in the rat striatum. Given that NO signaling potently modulates the membrane excitability of striatal projection neurons, it is plausible that up-regulation of NOS activity after DA D2 receptor blockade contributes to the therapeutic efficacy and/or motor side effects associated with antipsychotic drugs.ObjectivesThis study assessed the impact of DA D2 receptor activation on striatal NOS activity in vivo. Characterization of the dopaminergic regulation of striatal NO signaling will be relevant for understanding the mechanism(s) of action of antipsychotic drugs.Materials and methodsStriatal NO efflux, evoked via electrical stimulation of the substantia nigra (SN) or systemic administration of the DA D1 receptor agonist SKF 81297, was assessed in anesthetized rats using an NO-selective amperometric microsensor.ResultsThe facilitatory effect of SN stimulation on striatal NO efflux was attenuated by systemic administration of the DA D2 receptor agonist quinpirole. Conversely, administration of the DA D2 receptor antagonist eticlopride augmented evoked NO efflux. NO efflux induced by systemic administration of SKF 81297 was attenuated by quinpirole and restored by co-administration of quinpirole and eticlopride. The facilitatory effect of SKF 81297 on NO efflux was also significantly attenuated after pretreatment with the non-specific NOS inhibitor methylene blue.ConclusionsActivation of NO synthesis by phasic DA transmission is down-regulated via a DA D2 receptor-dependent mechanism. DA D2 receptor activation opposes DA D1 receptor activation of NO synthesis at a site postsynaptic to the DA terminal. Further studies examining NO–DA dynamics may have potential to reveal novel therapeutic strategies to treat various brain disorders.

Frontal cortical afferents facilitate striatal nitric oxide transmission in vivo via a NMDA receptor and neuronal NOS-dependent mechanism.

Striatal nitric oxide (NO) signaling plays a critical role in modulating neural processing and motor behavior. Nitrergic interneurons receive synaptic inputs from corticostriatal neurons and are activated via ionotropic glutamate receptor stimulation. However, the afferent regulation of NO signaling is poorly characterized. The role of frontal cortical afferents in regulating NO transmission was assessed in anesthetized rats using amperometric microsensor measurements of NO efflux and local field potential recordings. Low frequency (3 Hz) electrical stimulation of the ipsilateral cortex did not consistently evoke detectable changes in striatal NO efflux. In contrast, train stimulation (30 Hz) of frontal cortical afferents facilitated NO efflux in a stimulus intensity‐dependent manner. Nitric oxide efflux evoked by train stimulation was transient, reproducible over time, and attenuated by systemic administration of either the NMDA receptor antagonist MK‐801 or the neuronal NO synthase inhibitors 7‐nitroindazole and NG‐propyl‐l‐arginine. The interaction between NO efflux evoked via train stimulation and local striatal neuron activity was assessed using dual microsensor and local field potential recordings carried out concurrently in the contralateral and ipsilateral striatum, respectively. Systemic administration of the non‐specific NO synthase inhibitor methylene blue attenuated both evoked NO efflux and the peak oscillation frequency (within the delta band) of local field potentials recorded immediately after train stimulation. Taken together, these observations indicate that feed‐forward activation of neuronal NO signaling by phasic activation of frontal cortical afferents facilitates the synchronization of glutamate driven oscillations in striatal neurons. Thus, NO signaling may act to amplify coherent corticostriatal transmission and synchronize striatal output.

Impact of dopamine–glutamate interactions on striatal neuronal nitric oxide synthase activity

RationaleIt is known that dopamine (DA) D1 receptor activation stimulates striatal nitric oxide (NO) synthesis, whereas D2 receptor activation produces the opposite effect. However, the mechanisms involved in the dopaminergic modulation of nitric oxide synthase (NOS) are unknown.ObjectivesWe hypothesized that the effects of DA on striatal NO signaling are dependent on ongoing glutamatergic activation of NOS. Therefore, the current study examined whether intact N-methyl-d-aspartic acid (NMDA) receptor activation is required for the dopaminergic modulation of NOS activity.MethodsWe assessed the impact of pharmacological manipulations of D1, D2, and NMDA receptors on NOS activity in the dorsal striatum and motor cortex using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry. Drugs were administered systemically to conscious animals and NADPH-d staining was quantified in these regions using ex vivo measurements of tissue optical density.ResultsAdministration of the neuronal NOS inhibitor NG-propyl-l-arginine (NPA), the D1 receptor antagonist SCH 23390, and the NMDA receptor antagonist 3-phosphonopropyl-piperazine-2-carboxylic acid (CPP) all attenuated staining selectively in the striatum. Administration of the D2 receptor agonist quinpirole decreased NADPH-d staining in both the striatum and cortex. Striatal NADPH-d staining elicited by administration of the D1 receptor agonist SKF 81297 or the D2 receptor antagonist eticlopride was attenuated by NPA, SCH 23390, and CPP pretreatment. Quinpirole pretreatment also abolished the facilitatory effect of SKF 81297.ConclusionsThese studies show for the first time that ongoing NMDA receptor activation is necessary for the modulation of striatal NOS activity by both facilitatory (D1 receptor activation) and inhibitory (D2 receptor activation) dopaminergic signaling mechanisms.

Feed-forward excitation of striatal neuron activity by frontal cortical activation of nitric oxide signaling in vivo

The gaseous neurotransmitter nitric oxide plays an important role in the modulation of corticostriatal synaptic transmission. This study examined the impact of frontal cortex stimulation on striatal nitric oxide efflux and neuron activity in urethane‐anesthetized rats using amperometric microsensor and single‐unit extracellular recordings, respectively. Systemic administration of the neuronal nitric oxide synthase inhibitor 7‐nitroindazole decreased spontaneous spike activity without affecting activity evoked by single‐pulse stimulation of the ipsilateral cortex. Train (30 Hz) stimulation of the contralateral frontal cortex transiently increased nitric oxide efflux in a robust and reproducible manner. Evoked nitric oxide efflux was attenuated by systemic administration of 7‐nitroindazole and the non‐selective nitric oxide synthase inhibitor NG‐nitro‐l‐arginine methyl ester. Train stimulation of the contralateral cortex, in a manner identical to that used to evoke nitric oxide efflux, had variable effects on spike activity assessed during the train stimulation trial, but induced a short‐term depression of cortically evoked activity in the first post‐train stimulation trial. Interestingly, 7‐nitroindazole potently decreased cortically evoked activity recorded during the train stimulation trial. Moreover, the short‐term depression of spike activity induced by train stimulation was enhanced following pretreatment with 7‐nitroindazole and attenuated after systemic administration of the dopamine D2 receptor antagonist eticlopride. These results demonstrate that robust activation of frontal cortical afferents in the intact animal activates a powerful nitric oxide‐mediated feed‐forward excitation which partially offsets concurrent D2 receptor‐mediated short‐term inhibitory influences on striatal neuron activity. Thus, nitric oxide signaling is likely to play an important role in the integration of corticostriatal sensorimotor information in striatal networks.

Nitric oxide-soluble guanylyl cyclase signaling regulates corticostriatal transmission and short-term synaptic plasticity of striatal projection neurons recorded in vivo

Striatal medium-sized spiny neurons (MSNs) contain the highest levels of soluble guanylyl cyclase (sGC) in the brain. Striatal sGC signaling is activated by nitric oxide (NO) and other neuromodulators. MSNs also express cGMP-dependent protein kinase and other components of the cGMP signaling system which are critically involved in integrating corticostriatal transmission and regulating synaptic plasticity in striatal networks. However, the influence of tonic and phasic activation of this signaling pathway on striatal MSN activity is poorly understood. The present study examined the impact of systemic administration of the selective sGC inhibitor [1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one] (ODQ) on spike activity evoked using low and high frequency electrical stimulation of the frontal cortex. MSN activity was monitored using single-unit extracellular recordings in urethane-anesthetized rats. ODQ administration significantly decreased spike activity evoked by low frequency cortical stimulation in a stimulus intensity- and time-dependent manner. Additionally, ODQ administered along with the neuronal NO synthase inhibitor 7-nitroindazole (7-NI) potently decreased the incidence of excitatory responses observed during high-frequency train stimulation of the contralateral frontal cortex. The short-term depression of cortically-evoked spike activity induced by train stimulation was enhanced following pretreatment with ODQ in MSNs exhibiting an excitatory response during cortical train stimulation. Unexpectedly, this effect of ODQ was reversed in animals receiving co-administration of ODQ and 7-NI. 7-NI/ODQ co-administration also reversed measures of short-term depression observed in MSNs exhibiting an inhibitory response during cortical train stimulation. These observations extend previous studies showing that tonic and phasic NO-sGC signaling modulates the responsiveness of MSNs to corticostriatal input. Moreover, phasic activation of NO signaling is likely to regulate short-term changes in corticostriatal synaptic plasticity via complex mechanisms involving both sGC-cGMP-dependent and independent pathways.

Inhibition of striatal soluble guanylyl cyclase-cGMP signaling reverses basal ganglia dysfunction and akinesia in experimental parkinsonism.

Objective There is clearly a necessity to identify novel non-dopaminergic mechanisms as new therapeutic targets for Parkinsons disease (PD). Among these, the soluble guanylyl cyclase (sGC)-cGMP signaling cascade is emerging as a promising candidate for second messenger-based therapies for the amelioration of PD symptoms. In the present study, we examined the utility of the selective sGC inhibitor 1H-[1], [2], [4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ) for reversing basal ganglia dysfunction and akinesia in animal models of PD. Methods The utility of the selective sGC inhibitor ODQ for reversing biochemical, electrophysiological, histochemical, and behavioral correlates of experimental PD was performed in 6-OHDA-lesioned rats and mice chronically treated with MPTP. Results We found that one systemic administration of ODQ is sufficient to reverse the characteristic elevations in striatal cGMP levels, striatal output neuron activity, and metabolic activity in the subthalamic nucleus observed in 6-OHDA-lesioned rats. The latter outcome was reproduced after intrastriatal infusion of ODQ. Systemic administration of ODQ was also effective in improving deficits in forelimb akinesia induced by 6-OHDA and MPTP. Interpretation Pharmacological inhibition of the sGC-cGMP signaling pathway is a promising non-dopaminergic treatment strategy for restoring basal ganglia dysfunction and attenuating motor symptoms associated with PD.

Acute cocaine administration increases NO efflux in the rat prefrontal cortex via a neuronal NOS-dependent mechanism.

An understanding of the neurochemical changes occurring following exposure to psychostimulants such as cocaine is critical for the development of novel pharmacotherapies aimed at disrupting the addictive cycle. It is well established that the acute effects of cocaine associated with drug‐induced blockade of dopamine (DA) reuptake processes occur in reward‐related areas of the brain including the medial prefrontal cortex (mPFC). Considerable evidence has accumulated indicating that the interaction between DA, glutamate, and nitric oxide (NO) is likely to play a critical role in the neuroplastic changes associated with psychostimulant exposure. However, the potential impact of cocaine on NO synthase (NOS) activity in the mPFC has not been examined. In this study, NO efflux was measured in the mPFC of anesthetized male rats using a NO‐selective electrochemical microsensor. Acute systemic administration of cocaine significantly increased NO efflux in the mPFC in a time‐dependent manner. Similar injections using vehicle did not affect NO efflux. The facilitatory effect of cocaine on NO efflux was transient and reproducible. The signal was derived from neuronal sources of NO, because it was attenuated by systemic administration of the neuronal NO synthase inhibitor 7‐nitroindazole. These studies support a role for prefrontal cortical NO signaling in cocaine‐induced changes in neurotransmission in reward‐related circuits involved in addiction. Synapse 62:710–713, 2008.

Striatal Nitric Oxide–cGMP Signaling in an Animal Model of Parkinson’s Disease

Nitric oxide (NO) is an important, yet understudied, gaseous regulator of neuronal activity in corticostriatal circuits. Numerous findings indicate that striatal NO signaling plays a key role in the regulation of shortand long-term synaptic plasticity [1, 2], protein kinase and protein phosphatase activities [3, 4], and gene expression [5]. Thus, disruption of striatal NO signaling cascades results in profound changes in behavioral, electrophysiological, and molecular responses to pharmacological manipulations of dopamine (DA) and glutamate transmission [6–8]. Most studies indicate that NO signaling facilitates neuronal activity via the activation of guanylyl cyclase (GC) (see [9] for review). It has been proposed that neuroadaptations in the NO–GC signaling cascade induced by DA depletion have an important role in the pathophysiology of Parkinson’s disease (PD). We will review this evidence and describe our recent work using various in vivo electrophysiological and neuroanatomical techniques aimed at determining how unilateral partial DA lesions alter NO–GC signaling in striatal circuits. Our studies are beginning to unravel the complex neuroadaptations in NO–GC signaling cascades that emerge following modest depletion of striatal DA that potentially mimics the early stages of PD. These neuroadaptations are likely involved in the enduring changes in glutamatergic transmission and motor behaviors observed in parkinsonian animal models and patients with PD.