Interferon-γ Involvement in the Neuroinflammation Associated with Parkinson's Disease and L-DOPA-Induced Dyskinesia.

Interferon-γ (IFN-γ) is a proinflammatory cytokine that activates glial cells. IFN-γ is increased in the plasma and brain of Parkinson's disease patients, suggesting its potential role in the disease. We investigated whether the IFN-γ deficiency could interfere with nigrostriatal degeneration induced by the neurotoxin 6-hydroxydopamine, L-DOPA-induced dyskinesia, and the neuroinflammatory features as astrogliosis, microgliosis, and induced nitric oxide synthase (iNOS) immunoreactivity induced by L-DOPA treatment. Wild type (WT) and IFN-γ knockout (IFN-γ/KO) mice received unilateral striatal microinjections of 6-hydroxydopamine. Animals were sacrificed 1, 3, 7, and 21 days after lesions. Additional group of WT and IFN-γ/KO parkinsonian mice, after 3 weeks of neurotoxin injection, received L-DOPA (intraperitoneally, for 21 days) resulting in dyskinetic-like behavior. Tyrosine hydroxylase immunostaining indicated the starting of dopaminergic lesion since the first day past toxin administration, progressively increased until the third day when it stabilized. There was no difference in the lesion and L-DOPA-induced dyskinesia intensity between WT and IFN-γ/KO mice. Remarkably, IFN-γ/KO mice treated with L-DOPA presented in the lesioned striatum an increase of iNOS and glial fibrilary acid protein (GFAP) density, compared with the WT group. Morphological analysis revealed the rise of astrocytes and microglia reactivity in IFN-γ/KO mice exibiting dyskinesia. In conclusion, IFN-γ/KO mice presented an intensification of the inflammatory reaction accompanying L-DOPA treatment and suggest that iNOS and GFAP increase, and the activation of astrocytes and microglia induced afterward L-DOPA treatment was IFN-γ independent events. Intriguingly, IFN-γ absence did not affect the degeneration of dopaminergic neurons or LID development.

Nociceptive Response to L-DOPA-Induced Dyskinesia in Hemiparkinsonian Rats.

Non-motor symptoms are increasingly identified to present clinical and diagnostic importance for Parkinson's disease (PD). The multifactorial origin of pain in PD makes this symptom of great complexity. The dopamine precursor, L-DOPA (L-3,4-dihydroxyphenylalanine), the classic therapy for PD, seems to be effective in pain threshold; however, there are no studies correlating L-DOPA-induced dyskinesia (LID) and nociception development in experimental Parkinsonism. Here, we first investigated nociceptive responses in a 6-hydroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease to a hind paw-induced persistent inflammation. Further, the effect of L-DOPA on nociception behavior at different times of treatment was investigated. Pain threshold was determined using von Frey and Hot Plate/Tail Flick tests. Dyskinesia was measured by abnormal involuntary movements (AIMs) induced by L-DOPA administration. This data is consistent to show that 6-OHDA-lesioned rats had reduced nociceptive thresholds compared to non-lesioned rats. Additionally, when these rats were exposed to a persistent inflammatory challenge, we observed increased hypernociceptive responses, namely hyperalgesia. L-DOPA treatment alleviated pain responses on days 1 and 7 of treatment, but not on day 15. During that period, we observed an inverse relationship between LID and nociception threshold in these rats, with a high LID rate corresponding to a reduced nociception threshold. Interestingly, pain responses resulting from CFA-induced inflammation were significantly enhanced during established dyskinesia. These data suggest a pro-algesic effect of L-DOPA-induced dyskinesia, which is confirmed by the correlation founded here between AIMs and nociceptive indexes. In conclusion, our results are consistent with the notion that central dopaminergic mechanism is directly involved in nociceptive responses in Parkinsonism condition.

Mice lacking interleukin-18 gene display behavioral changes in animal models of psychiatric disorders: Possible involvement of immunological mechanisms.

Preclinical and clinical evidence suggests pro-inflammatory cytokines might play an important role in the neurobiology of schizophrenia and stress-related psychiatric disorders. Interleukin-18 (IL-18) is a member of the IL-1 family of cytokines and it is widely expressed in brain regions involved in emotional regulation. Since IL-18 involvement in the neurobiology of mental illnesses, including schizophrenia, remains unknown, this work aimed at investigating the behavior of IL-18 null mice (KO) in different preclinical models: 1. the prepulse inhibition test (PPI), which provides an operational measure of sensorimotor gating and schizophrenic-like phenotypes; 2. amphetamine-induced hyperlocomotion, a model predictive of antipsychotic activity; 3. resident-intruder test, a model predictive of aggressive behavior. Furthermore, the animals were submitted to models used to assess depressive- and anxiety-like behavior. IL-18KO mice showed impaired baseline PPI response, which was attenuated by d-amphetamine at a dose that did not modify PPI response in wild-type (WT) mice, suggesting a hypodopaminergic prefrontal cortex function in those mice. d-Amphetamine, however, induced hyperlocomotion in IL-18KO mice compared to their WT counterparts, suggesting hyperdopaminergic activity in the midbrain. Moreover, IL-18KO mice presented increased basal levels of IL-1ß levels in the hippocampus and TNF-α in the prefrontal cortex, suggesting an overcompensation of IL-18 absence by increased levels of other proinflammatory cytokines. Although no alteration was observed in the forced swimming or in the elevated plus maze tests in naïve IL-18KO mice, these mice presented anxiogenic-like behavior after exposure to repeated forced swimming stress. In conclusion, deletion of the IL-18 gene resembled features similar to symptoms observed in schizophrenia (positive and cognitive symptoms, aggressive behavior), in addition to increased susceptibility to stress. The IL-18KO model, therefore, could provide new insights into how changes in brain immunological homeostasis induce behavioral changes related to psychiatric disorders, such as schizophrenia.

Cannabidiol effects in the prepulse inhibition disruption induced by amphetamine.

RATIONALE: The information processing appears to be deficient in schizophrenia. Prepulse inhibition (PPI), which measures the inhibition of a motor response by a weak sensory event, is considered particularly useful to understand the biology of information processing in schizophrenia patients. Drugs that facilitate dopaminergic neurotransmission such as amphetamine induce PPI disruption in human and rodents. Clinical and neurobiological findings suggest that the endocannabinoid system and cannabinoids may be implicated in the pathophysiology and treatment of schizophrenia. Cannabidiol (CBD), a non-psychotomimetic constituent of the Cannabis sativa plant, has also been reported to have potential as an antipsychotic. OBJECTIVE: Our aim was to investigate if CBD pretreatment was able to prevent PPI disruption induced by amphetamine. Since one possible mechanism of CBD action is the facilitation of endocannabinoid-mediated neurotransmission through anandamide, we tested the effects of an anandamide hydrolysis inhibitor (URB597) in the amphetamine-induced PPI disruption. METHODS: Male Swiss mice were treated with CBD systemic or intra-accumbens, or URB597 (systemic) prior to amphetamine and were exposed to PPI test. RESULTS: Amphetamine (10 mg/kg) disrupted PPI while CBD (15-60 mg/kg) or URB597 (0.1-1 mg/kg) administered alone had no effect. Pretreatment with CBD attenuated the amphetamine-disruptive effects on PPI test after systemic or intra-accumbens administration. Similar effects were also found with the inhibitor of anandamide hydrolysis. CONCLUSION: These results corroborate findings indicating that CBD induces antipsychotic-like effects. In addition, they pointed to the nucleus accumbens as a possible site of these effects. The increase of anandamide availability may be enrolled in the CBD effects.

Descending mechanisms activated by the anterior pretectal nucleus initiate but do not maintain neuropathic pain in rats.

BACKGROUND: The anterior pretectal nucleus (APtN) activates descending mechanisms of pain control. This study evaluated whether the APtN also controls neuropathic pain in rats. METHODS: The hypersensitivity to mechanical stimulation with an electronic von Frey apparatus and the number of Fos-immunoreactive (Fos-ir) neurons in the APtN were evaluated in rats before and after chronic constriction injury of the sciatic nerve. RESULTS: The tactile hypersensitivity was characterized by an initial phase (the 2 days following the injury) and a maintenance phase (the subsequent 7 days). The injection of 2% lidocaine (0.25 µL) or N-methyl-D-aspartate (2.5 µg/0.25 µL) into the APtN intensified the tactile hypersensitivity observed 2 days after injury but did not alter the tactile hypersensitivity observed 7 and 14 days after injury. The injection of naloxone (10 ng/0.25 µL) or methysergide (40 pg/0.25 µL) but not atropine (100 ng/0.25 µL) into the APtN also intensified the tactile hypersensitivity observed 2 days after the injury. A significant increase in the number of Fos-ir cells was found in the contralateral APtN 2 days but not 7 or 14 days after the injury. Electrical stimulation of the APtN reduced the tactile hypersensitivity at 2, 7 and 14 days after the nerve ligation. CONCLUSION: APtN exerts a tonic inhibitory influence on persistent pain. The results point out to an important role of opioid and serotonergic mediation into the APtN to inhibit hyperalgesia during the initial phase of neuropathic pain.

7-Nitroindazole blocks the prepulse inhibition disruption and c-Fos increase induced by methylphenidate.

RATIONALE: The dopamine and nitric oxide (NO) interaction on sensorimotor gating modulation measured through the prepulse inhibition (PPI), has been described recently. The PPI impairment has been reported in several neuropsychiatric conditions, particularly in schizophrenia. We previously demonstrated that NO inhibitors, similarly to the antipsychotic drugs, attenuate the disruptive effect of amphetamine or its analogue methylphenidate in the PPI response. OBJECTIVES: Our aim was to determine if the known expression of the neuronal activity marker c-Fos induced by methylphenidate may be modified by NO inhibition. Mice were treated with the PPI-disruptive dose of methylphenidate (30 mg/kg) preceded by pretreatment with saline, or the dose of preferential neuronal NO inhibitor 7-Nitroindazole (7NI; 10 mg/kg) which promotes PPI recovery. RESULTS: Acute treatment with methylphenidate at dose that caused PPI disruption induced a robust increase in the number of c-Fos-positive cells in the cingulate and motor cortex, dorsal, dorsolateral, and ventrolateral striatum, nucleus accumbens core and shell, and basolateral amygdala. In the animals which presented PPI recovery through 7NI pretreatment, the c-Fos increase induced by methylphenidate was significantly reduced in the cingulate cortex (rostral level), striatum, mainly dorsal and ventrolateral, nucleus accumbens (core and shell), and in the basolateral amygdala. CONCLUSION: Our results suggest that 7NI effects appear to be related to its ability to prevent the activation of specific brain areas, including nucleus accumbens and amygdala, counteracting the stimulant effects of methylphenidate in these regions.

Hippocampal ether-à-go-go1 potassium channels blockade: effects in the startle reflex and prepulse inhibition.

Recently, our group described the ether-à-go-go1(Eag1) voltage-gated potassium (K(+)) channel (Kv10.1) expression in the dopaminergic cells indicating that these channels are part of the diversified group of ion channels related to dopaminergic neurons function. The increase of dopamine neurotransmission induces a reduction in the prepulse inhibition (PPI) of the acoustic startle reflex in rodents, which is a reliable index of sensorimotor gating deficits. The PPI response has been reported to be abnormally reduced in schizophrenia patients. The role of Eag1 K(+) channels in the PPI reaction had not been revealed until now, albeit the singular distribution of Eag1 in the dentate gyrus of the hippocampus and the hippocampal regulation of the startle reflex and PPI. The aim of this work was to investigate if Eag1 blockade on hippocampus modifies the PPI-disruptive effects of apomorphine in Wistar rats. Bilateral injection of anti-Eag1 single-chain antibody into the dentate gyrus of hippocampus did not modify apomorphine-disruptive effects in the PPI response. However, Eag1 antibody completely restored the startle amplitude decrease revealed after dentate gyrus surgery. These potentially biological important phenomenon merits further investigation regarding the role of Eag1 K(+) channels, mainly, on startle reflex modulation, since the physiological role of these channels remain obscure.

Effect of melatonin on the functional recovery from experimental traumatic compression of the spinal cord.

Spinal cord injury is an extremely severe condition with no available effective therapies. We examined the effect of melatonin on traumatic compression of the spinal cord. Sixty male adult Wistar rats were divided into three groups: sham-operated animals and animals with 35 and 50% spinal cord compression with a polycarbonate rod spacer. Each group was divided into two subgroups, each receiving an injection of vehicle or melatonin (2.5 mg/kg, intraperitoneal) 5 min prior to and 1, 2, 3, and 4 h after injury. Functional recovery was monitored weekly by the open-field test, the Basso, Beattie and Bresnahan locomotor scale and the inclined plane test. Histological changes of the spinal cord were examined 35 days after injury. Motor scores were progressively lower as spacer size increased according to the motor scale and inclined plane test evaluation at all times of assessment. The results of the two tests were correlated. The open-field test presented similar results with a less pronounced difference between the 35 and 50% compression groups. The injured groups presented functional recovery that was more evident in the first and second weeks. Animals receiving melatonin treatment presented more pronounced functional recovery than vehicle-treated animals as measured by the motor scale or inclined plane. NADPH-d histochemistry revealed integrity of the spinal cord thoracic segment in sham-operated animals and confirmed the severity of the lesion after spinal cord narrowing. The results obtained after experimental compression of the spinal cord support the hypothesis that melatonin may be considered for use in clinical practice because of its protective effect on the secondary wave of neuronal death following the primary wave after spinal cord injury.

Investigations into Potential Extrasynaptic Communication between the Dopaminergic and Nitrergic Systems.

Nitric oxide is unconstrained by cell membranes and can therefore act along a broad distance as a volume transmitter. Spillover of nitric oxide between neurons may have a major impact on central nervous system diseases and particularly on neurodegeneration. There is evidence whereby communication between nitrergic and dopaminergic systems plays an essential role in the control of the nigrostriatal pathway. However, there is sparse information for either the coexistence or overlap of nitric oxide and dopaminergic structures. The dual localization of immunoreactivity for nitric oxide synthase (NOS) and tyrosine hydroxylase, enzymes responsible for the synthesis of nitric oxide and dopamine, respectively, was examined in neurons of the nigrostriatal pathway in the rat brain by means of a double-immunohistochemical method and confocal laser scanning microscopy, acquired at the resolution limit. After perfusional fixation, the brains were cut and double-immunostained. A proximity analysis of tyrosine hydroxylase and NOS structures was done using binary masks generated from the respective maximum projections, using confocal laser microscopy. Unrevealed regions were determined somatodendritic positive for both NOS and tyrosine hydroxylase, within an image limit resolution at 2 µm-wide margin. The described interconnected localization of nNOS(+) and TH(+) containing neuronal fibers and cells bodies in the nigrostriatal pathway propose a close anatomical link between the two neurotransmitters.

Eag1, Eag2, and SK3 potassium channel expression in the rat hippocampus after global transient brain ischemia.

Transient global brain ischemia causes delayed neuronal death in the hippocampus that has been associated with impairments in hippocampus-dependent brain function, such as mood, learning, and memory. We investigated the expression of voltage-dependent Kcnh1 and Kcnh5, ether à go-go-related Eag1 and Eag2 (K(V) 10.1 and K(V) 10.2), and small-conductance calcium-activated SK3 (K(Ca) 2.3, Kcnn3) K(+) channels in the hippocampus in rats after transient global brain ischemia. We tested whether the expression of these channels is associated with behavioral changes by evaluating the animals in the elevated plus maze and step-down inhibitory avoidance task. Seven or tweny-eight days after transient global brain ischemia, one group of rats had the hippocampus bilaterally dissected, and mRNA levels were determined. Seven days after transient global brain ischemia, the rats exhibited a decrease in anxiety-like behavior and memory impairments. An increase in anxiety levels was detected 28 days after ischemia. Eag2 mRNA downregulation was observed in the hippocampus 7 days after transient global brain ischemia, whereas Eag1 and SK3 mRNA expression remained unaltered. This is the first experimental evidence that transient global brain ischemia temporarily alters Eag2. The number of intact-appearing pyramidal neurons was substantially decreased in CA1 and statistically measurable in CA2, CA3, and CA4 hippocampal subfields compared with sham control animals 7 or 28 days after ischemia. mRNA expression in the rat hippocampus. The present results provide further information for the characterization of the physiological role of Eag2 channels in the central nervous system.

Ether-à-go-go 1 (Eag1) potassium channel expression in dopaminergic neurons of basal ganglia is modulated by 6-hydroxydopamine lesion.

The ether à go-go (Eag) gene encodes the voltage-gated potassium (K(+)) ion channel Kv10.1, whose function still remains unknown. As dopamine may directly affect K(+) channels, we evaluated whether a nigrostriatal dopaminergic lesion induced by the neurotoxin 6-hydroxydopamine (6-OHDA) would alter Eag1-K(+) channel expression in the rat basal ganglia and related brain regions. Male Wistar rats received a microinjection of either saline or 6-OHDA (unilaterally) into the medial forebrain bundle. The extent of the dopaminergic lesion induced by 6-OHDA was evaluated by apomorphine-induced rotational behavior and by tyrosine hydroxylase (TH) immunoreactivity. The 6-OHDA microinjection caused a partial or complete lesion of dopaminergic cells, as well as a reduction of Eag1+ cells in a manner proportional to the extent of the lesion. In addition, we observed a decrease in TH immunoreactivity in the ipsilateral striatum. In conclusion, the expression of the Eag1-K(+)-channel throughout the nigrostriatal pathway in the rat brain, its co-localization with dopaminergic cells and its reduction mirroring the extent of the lesion highlight a physiological circuitry where the functional role of this channel can be investigated. The Eag1-K(+) channel expression in dopaminergic cells suggests that these channels are part of the diversified group of ion channels that generate and maintain the electrophysiological activity pattern of dopaminergic midbrain neurons.

Nitric oxide synthase inhibitors improve prepulse inhibition responses of Wistar rats.

INTRODUCTION: Cognitive and attentional deficits in schizophrenia include impairment of the sensorimotor filter as measured by prepulse inhibition (PPI). In this way, the study of animals that naturally present low PPI responses could be a useful approach for screening new antipsychotic drugs. Several pieces of evidence suggest that dopamine and nitric oxide (NO) can modulate PPI but their role in those animals is unknown. OBJECTIVES: The aim of this study was to investigate the role of dopamine and NO in Wistar rats with naturally low PPI response. METHODS: Male Wistar rats with low PPI responses received an i.p. injection of the antipsychotics haloperidol (0.1, 0.3 or 1mg/kg) or clozapine (0.5, 1.5 or 5mg/kg), the anxiolytic diazepam (1 or 3mg/kg) or the NO synthase (NOS) inhibitors, N(G)- nitro-l-arginine (l-NOARG; 40mg/kg, acutely or sub-chronically) or 7-Nitroindazole (7-NI; 3, 10 or 30mg/kg). All animals were submitted to the PPI test 1h after injection. Striatal and cortical dopamine, DOPAC, and noradrenaline levels of rats with low PPI responses were compared to rats with normal PPI responses. RESULTS: We found increased levels of catecholamines on the striatum and prefrontal cortex of Wistar rats with low PPI. In these animals, both antipsychotics, typical and atypical, and NOS inhibitors significantly increased PPI. CONCLUSION: Taken together, our findings suggest that the low PPI phenotype may be driven by an overactive catecholamine system. Additionally, our results corroborate the hypothesis of dopamine and NO interaction on PPI modulation and suggest that Wistar rats with low PPI may represent an interesting non-pharmacological model to evaluate new potential antipsychotics.

Lack of tolerance for the anti-dyskinetic effects of 7-nitroindazole, a neuronal nitric oxide synthase inhibitor, in rats.

7-Nitroindazole (7-NI) inhibits neuronal nitric oxide synthase in vivo and reduces l-DOPA-induced dyskinesias in a rat model of parkinsonism. The aim of the present study was to determine if the anti-dyskinetic effect of 7-NI was subject to tolerance after repeated treatment and if this drug could interfere with the priming effect of l-DOPA. Adult male Wistar rats (200-250 g) with unilateral depletion of dopamine in the substantia nigra compacta were treated with l-DOPA (30 mg/kg) for 34 days. On the 1st day, 6 rats received ip saline and 6 received ip 7-NI (30 mg/kg) before l-DOPA. From the 2nd to the 26th day, all rats received l-DOPA daily and, from the 27th to the 34th day, they also received 7-NI before l-DOPA. Animals were evaluated before the drug and 1 h after l-DOPA using an abnormal involuntary movement scale and a stepping test. All rats had a similar initial motor deficit. 7-NI decreased abnormal involuntary movement induced by l-DOPA and the effect was maintained during the experiment before 7-NI, median (interquartile interval), day 26: 16.75 (15.88-17.00); day 28: 0.00 (0.00-9.63); day 29: 13.75 (2.25-15.50); day 30: 0.5 (0.00-6.25); day 31: 4.00 (0.00-7.13), and day 34: 0.5 (0.00-14.63), Friedman followed by Wilcoxon test,vs day 26, P < 0.05;. The response to l-DOPA alone was not modified by the use of 7-NI before the first administration of the drug (l-DOPA vs time interaction, F1,10 = 1.5, NS). The data suggest that tolerance to the anti-dyskinetic effects of a neuronal nitric oxide synthase inhibitor does not develop over a short-term period of repeated administration. These observations open a possible new therapeutic approach to motor complications of chronic l-DOPA therapy in patients with Parkinson's disease.

Lack of tolerance for the anti-dyskinetic effects of 7-nitroindazole, a neuronal nitric oxide synthase inhibitor, in rats

7-Nitroindazole (7-NI) inhibits neuronal nitric oxide synthase in vivo and reduces l-DOPA-induced dyskinesias in a rat model of parkinsonism. The aim of the present study was to determine if the anti-dyskinetic effect of 7-NI was subject to tolerance after repeated treatment and if this drug could interfere with the priming effect of l-DOPA. Adult male Wistar rats (200-250 g) with unilateral depletion of dopamine in the substantia nigra compacta were treated with l-DOPA (30 mg/kg) for 34 days. On the 1st day, 6 rats received ip saline and 6 received ip 7-NI (30 mg/kg) before l-DOPA. From the 2nd to the 26th day, all rats received l-DOPA daily and, from the 27th to the 34th day, they also received 7-NI before l-DOPA. Animals were evaluated before the drug and 1 h after l-DOPA using an abnormal involuntary movement scale and a stepping test. All rats had a similar initial motor deficit. 7-NI decreased abnormal involuntary movement induced by l-DOPA and the effect was maintained during the experiment before 7-NI, median (interquartile interval), day 26: 16.75 (15.88-17.00); day 28: 0.00 (0.00-9.63); day 29: 13.75 (2.25-15.50); day 30: 0.5 (0.00-6.25); day 31: 4.00 (0.00-7.13), and day 34: 0.5 (0.00-14.63), Friedman followed by Wilcoxon test,vs day 26, P < 0.05;. The response to l-DOPA alone was not modified by the use of 7-NI before the first administration of the drug (l-DOPA vs time interaction, F1,10 = 1.5, NS). The data suggest that tolerance to the anti-dyskinetic effects of a neuronal nitric oxide synthase inhibitor does not develop over a short-term period of repeated administration. These observations open a possible new therapeutic approach to motor complications of chronic l-DOPA therapy in patients with Parkinson’s disease.

Tolerance to the cataleptic effect that follows repeated nitric oxide synthase inhibition may be related to functional enzymatic recovery.

Systemic or intra-striatal acute administration of nitric oxide synthase (NOS) inhibitors causes catalepsy in rodents. This effect disappears after sub-chronic treatment. The aim of the present study was to investigate if this tolerance is related to changes in the expression of NOS or dopamine-2 (D2) receptor or to a recovery of NOS activity. Male albino Swiss mice (25-30 g) received single or sub-chronic (once a day for 4 days) i.p. injections of saline or L-nitro-arginine (L-NOARG, 40 mg/kg), a non-selective inhibitor of neuronal nitric oxide synthase (nNOS). Twenty-four hours after the last injection, the animals were killed and their brains were removed for immunohistochemistry assay to detect the presence of nNOS or for 'in-situ' hybridisation study using (35)S-labeled oligonucleotide probe complementary to D2 receptor mRNA. The results were analysed by computerised densitometry. Independent groups of animals received the same treatment, but were submitted to the catalepsy test and had their brain removed to measure nitrite and nitrate (NOx) concentrations in the striatum. Acute administration of L-NOARG caused catalepsy that disappeared after sub-chronic treatment. The levels of NOx were significantly reduced after acute L-NOARG treatment. The decrease in NOx after drug injection suffered a partial tolerance after sub-chronic treatment. The catalepsy time after acute or sub-chronic treatment with L-NOARG was negatively (r = -0.717) correlated with NOx levels. Animals that received repeated L-NOARG injections also showed an increase in the number of nNOS-positive neurons in the striatum. No change in D2 receptor mRNA expression was found in the dorsal striatum, nucleus accumbens and substantia nigra. Together, these results suggest that tolerance to L-NOARG cataleptic effects do not depend on changes in D2 receptors. They may depend, however, on plastic changes in nNOS neurons resulting in partial recovery of NO formation in the striatum.

Nitric oxide modulation of methylphenidate-induced disruption of prepulse inhibition in Swiss mice.

Drugs that facilitate dopaminergic neurotransmission induce cognitive and attentional deficits which include inability to filter sensory input measured by prepulse inhibition (PPI). Methylphenidate, an amphetamine analog is used in the treatment of attention deficit hyperactivity disorder. Given that nitric oxide (NO) modulates dopamine effect our aim is to analyze the nitric oxide synthase (NOS) and soluble guanylate cyclase (sGC) inhibitors effect on PPI disruption induced by methylphenidate. The inhibitors effects were compared to those produced by haloperidol and clozapine. Male Swiss mice received a first i.p. injection (one hour before testing), of either saline, or N(G) nitro l-arginine (10, 40 or 90 mg/kg), or 7-Nitroindazole (3, 10, 30 or 60 mg/kg), or oxadiazolo-quinoxalin (5 or 10 mg/kg), or haloperidol (1 mg/kg), or clozapine (5 mg/kg). Thirty min later mice received the second injection of either saline or methylphenidate (20 or 30 mg/kg) or amphetamine (5 or 10 mg/kg). One group of mice received intracerebroventricular 7-Nitroindazole (50 or 100 nM) followed by systemic administration of saline or methylphenidate (30 mg/kg). The results revealed a methylphenidate dose-dependent disruption of PPI comparable to amphetamine. The effect was prevented by either nitric oxide synthase or guanilate cyclase inhibitors or clozapine or haloperidol. In conclusion, methylphenidate induced a dose-dependent PPI disruption in Swiss mice modulated by dopamine and NO/sGC. The results corroborate the hypothesis of dopamine and NO interacting to modulate sensorimotor gating through central nervous system. It may be useful to understand methylphenidate and other psychostimulants effects.

Nitric oxide synthase inhibition attenuates L-DOPA-induced dyskinesias in a rodent model of Parkinson's disease.

Chronic L-DOPA pharmacotherapy in Parkinson's disease is often accompanied by the development of abnormal and excessive movements known as L-DOPA-induced dyskinesia. Rats with 6-hydroxydopamine lesion of dopaminergic neurons chronically treated with L-DOPA develop a rodent analog of this dyskinesia characterized by severe axial, limb, locomotor and orofacial abnormal involuntary movements. While the mechanisms by which these effects occur are not clear, they may involve the nitric oxide system. In the present study we investigate if nitric oxide synthase inhibitors can prevent dyskinesias induced by repeated administration of L-DOPA in rats with unilateral 6-hydroxydopamine lesion. Chronic L-DOPA (high fixed dose, 100 mg/kg; low escalating dose, 10-30 mg/kg) treatment induced progressive dyskinesia changes. Two nitric oxide synthase inhibitors, 7-nitroindazole (1-30 mg/kg) and NG-nitro-L-arginine (50 mg/kg), given 30 min before L-DOPA, attenuate dyskinesia. 7-Nitroindazolee also improved motor performance of these animals in the rota-rod test. These results suggest the possibility that nitric oxide synthase inhibitors may be useful to treat L-DOPA-induced dyskinesia.

Different role of isoproterenol and NOS inhibitors on salivary ducts of rats.

OBJECTIVES: Nitric oxide (NO) is a diffusible intracellular messenger that is present in saliva. Chronic treatment with isoproterenol, a beta receptor agonist, stimulates the release of NO from acinar cells and induces salivary gland hypertrophy. The aim of this study was to investigate the effect of NO synthesis inhibitors and isoproterenol on rat salivary glands. We analyzed salivary gland weight and the number of ducts per unit area (0.5mm(2)) by NADPH-diaphorase histochemistry (to identify the presence of the enzyme NO synthase-NOS) and haematoxylin-and-eosin (HE). METHODS: For 8 days male Wistar rats received daily single intraperitoneal injections of saline or a NOS inhibitor (40mg/kg N(omega)-nitro-L-arginine L-NOARG or N(omega)-nitro-l-arginine methyl ester L-NAME). This was followed, 30min later, by subcutaneous injection of isoproterenol (2 or 5mg/kg) or saline. RESULTS: Isoproterenol increased parotid and submandibular gland weights. Isoproterenol (2mg/kg) induced a decrease of ducts per unit area inversely correlated to the weight of the parotid gland. This effect was augmented by L-NAME. In the submandibular gland L-NAME attenuated isoproterenol (2mg/kg) weight increase. In the submandibular gland isoproterenol and NOS inhibitors induced an increase in ducts per unit area (HE and NADPH-diaphorase). No effect was observed in the sublingual gland. CONCLUSION: To our knowledge this is the first description of isoproterenol and NOS inhibitors increasing duct density in the submandibular gland. Our results corroborate the hypothesis that NO plays different roles in parotid and submandibular glands.

Nitric oxide synthase activity in tissues of the blowfly Chrysomya megacephala (Fabricius, 1794).

Although insects lack the adaptive immune response of the mammalians, they manifest effective innate immune responses, which include both cellular and humoral components. Cellular responses are mediated by hemocytes, and humoral responses include the activation of proteolytic cascades that initiate many events, including NO production. In mammals, nitric oxide synthases (NOSs) are also present in the endothelium, the brain, the adrenal glands, and the platelets. Studies on the distribution of NO-producing systems in invertebrates have revealed functional similarities between NOS in this group and vertebrates. We attempted to localize NOS activity in tissues of naïve (UIL), yeast-injected (YIL), and saline-injected (SIL) larvae of the blowfly Chrysomya megacephala, using the NADPH diaphorase technique. Our findings revealed similar levels of NOS activity in muscle, fat body, Malpighian tubule, gut, and brain, suggesting that NO synthesis may not be involved in the immune response of these larval systems. These results were compared to many studies that recorded the involvement of NO in various physiological functions of insects.

Nitric oxide synthase activity in tissues of the blowfly Chrysomya megacephala (Fabricius, 1794)

Although insects lack the adaptive immune response of the mammalians, they manifest effective innate immune responses, which include both cellular and humoral components. Cellular responses are mediated by hemocytes, and humoral responses include the activation of proteolytic cascades that initiate many events, including NO production. In mammals, nitric oxide synthases (NOSs) are also present in the endothelium, the brain, the adrenal glands, and the platelets. Studies on the distribution of NO-producing systems in invertebrates have revealed functional similarities between NOS in this group and vertebrates. We attempted to localize NOS activity in tissues of naïve (UIL), yeast-injected (YIL), and saline-injected (SIL) larvae of the blowfly Chrysomya megacephala, using the NADPH diaphorase technique. Our findings revealed similar levels of NOS activity in muscle, fat body, Malpighian tubule, gut, and brain, suggesting that NO synthesis may not be involved in the immune response of these larval systems. These results were compared to many studies that recorded the involvement of NO in various physiological functions of insects.