Ana Carolina Issy

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.

Cannabidiol Attenuates Sensorimotor Gating Disruption and Molecular Changes Induced by Chronic Antagonism of NMDA receptors in Mice

Background: Preclinical and clinical data suggest that cannabidiol (CBD), a major non-psychotomimetic compound from Cannabis sativa, induces antipsychotic-like effects. However, the antipsychotic properties of repeated CBD treatment have been poorly investigated. Behavioral changes induced by repeated treatment with glutamate N-methyl-D-aspartate receptor (NMDAR) antagonists have been proposed as an animal model of schizophrenia-like signs. In the present study, we evaluated if repeated treatment with CBD would attenuate the behavioral and molecular modifications induced by chronic administration of one of these antagonists, MK-801. Methods: Male C57BL/6J mice received daily i.p. injections of MK-801 (0.1, 0.5, or 1mg/kg) for 14, 21, or 28 days. Twenty-four hours after the last injection, animals were submitted to the prepulse inhibition (PPI) test. After that, we investigated if repeated treatment with CBD (15, 30, and 60mg/kg) would attenuate the PPI impairment induced by chronic treatment with MK-801 (1mg/kg; 28 days). CBD treatment began on the 6th day after the start of MK-801 administration and continued until the end of the treatment. Immediately after the PPI, the mice brains were removed and processed to evaluate the molecular changes. We measured changes in FosB/ΔFosB and parvalbumin (PV) expression, a marker of neuronal activity and a calcium-binding protein expressed in a subclass of GABAergic interneurons, respectively. Changes in mRNA expression of the NMDAR GluN1 subunit gene (GRN1) were also evaluated. CBD effects were compared to those induced by the atypical antipsychotic clozapine. Results: MK-801 administration at the dose of 1mg/kg for 28 days impaired PPI responses. Chronic treatment with CBD (30 and 60mg/kg) attenuated PPI impairment. MK-801 treatment increased FosB/ΔFosB expression and decreased PV expression in the medial prefrontal cortex. A decreased mRNA level of GRN1 in the hippocampus was also observed. All the molecular changes were attenuated by CBD. CBD by itself did not induce any effect. Moreover, CBD effects were similar to those induced by repeated clozapine treatment. Conclusions: These results indicate that repeated treatment with CBD, similar to clozapine, reverses the psychotomimetic-like effects and attenuates molecular changes observed after chronic administration of an NMDAR antagonist. These data support the view that CBD may have antipsychotic properties.

Experimental model of intervertebral disc degeneration by needle puncture in Wistar rats

Animal models of intervertebral disc degeneration play an important role in clarifying the physiopathological mechanisms and testing novel therapeutic strategies. The objective of the present study is to describe a simple animal model of disc degeneration involving Wistar rats to be used for research studies. Disc degeneration was confirmed and classified by radiography, magnetic resonance and histological evaluation. Adult male Wistar rats were anesthetized and submitted to percutaneous disc puncture with a 20-gauge needle on levels 6-7 and 8-9 of the coccygeal vertebrae. The needle was inserted into the discs guided by fluoroscopy and its tip was positioned crossing the nucleus pulposus up to the contralateral annulus fibrosus, rotated 360° twice, and held for 30 s. To grade the severity of intervertebral disc degeneration, we measured the intervertebral disc height from radiographic images 7 and 30 days after the injury, and the signal intensity T2-weighted magnetic resonance imaging. Histological analysis was performed with hematoxylin-eosin and collagen fiber orientation using picrosirius red staining and polarized light microscopy. Imaging and histological score analyses revealed significant disc degeneration both 7 and 30 days after the lesion, without deaths or systemic complications. Interobserver histological evaluation showed significant agreement. There was a significant positive correlation between histological score and intervertebral disc height 7 and 30 days after the lesion. We conclude that the tail disc puncture method using Wistar rats is a simple, cost-effective and reproducible model for inducing disc degeneration.

Critical role of nitric oxide in the modulation of prepulse inhibition in Swiss mice

RationaleNitric oxide (NO) modulates the dopamine uptake and release processes and appears to be implicated in dopamine-related pathologies, such as schizophrenia. However, it is unclear whether there is excess or deficient NO synthesis in schizophrenia pathophysiology. Analyses of the intracellular pathways downstream of NO system activation have identified the cyclic nucleotide cyclic guanosine monophosphate (cGMP) as a possible target for drug development. Defects in the sensorimotor gating of the neural mechanism underlying the integration and processing of sensory information have been detected across species through prepulse inhibition (PPI).ObjectivesThe aim of this study was to investigate the effects of NO/cGMP increase on sensorimotor gating modulation during dopamine hyperfunction.MethodsMice were treated with NO donors and subjected to the PPI test. Treatment with the NO donor sodium nitroprusside was preceded by pretreatment with a soluble guanylate cyclase (sGC) inhibitor. Additionally, the mice were treated with NO donors and phosphodiesterases inhibitors prior to amphetamine treatment.ResultsPretreatment with the NO donors enhanced the PPI response and attenuated the amphetamine-disruptive effects on the PPI. The sGC inhibitor did not modify the sodium nitroprusside effects. Additionally, the cGMP increase induced by a specific phosphodiesterase inhibitor did not modify the amphetamine-disruptive effect.ConclusionsThis study provides the first demonstration that an increase in NO can improve the PPI response and block the amphetamine-disruptive effects on the PPI response. Our data are consistent with recent clinical results. However, these effects do not appear to be related to an increase in cGMP levels, and further investigation is thus required.

Protective effects of cannabidiol on lesion-induced intervertebral disc degeneration.

Disc degeneration is a multifactorial process that involves hypoxia, inflammation, neoinnervation, accelerated catabolism, and reduction in water and glycosaminoglycan content. Cannabidiol is the main non-psychotropic component of the Cannabis sativa with protective and anti-inflammatory properties. However, possible therapeutic effects of cannabidiol on intervertebral disc degeneration have not been investigated yet. The present study investigated the effects of cannabidiol intradiscal injection in the coccygeal intervertebral disc degeneration induced by the needle puncture model using magnetic resonance imaging (MRI) and histological analyses. Disc injury was induced in the tail of male Wistar rats via a single needle puncture. The discs selected for injury were punctured percutaneously using a 21-gauge needle. MRI and histological evaluation were employed to assess the results. The effects of intradiscal injection of cannabidiol (30, 60 or 120 nmol) injected immediately after lesion were analyzed acutely (2 days) by MRI. The experimental group that received cannabidiol 120 nmol was resubmitted to MRI examination and then to histological analyses 15 days after lesion/cannabidiol injection. The needle puncture produced a significant disc injury detected both by MRI and histological analyses. Cannabidiol significantly attenuated the effects of disc injury induced by the needle puncture. Considering that cannabidiol presents an extremely safe profile and is currently being used clinically, these results suggest that this compound could be useful in the treatment of intervertebral disc degeneration.

Nitric oxide modulates dopaminergic regulation of prepulse inhibition in the basolateral amygdala

Systemic injection of the nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine (LNO) prevents the disruptive effect of amphetamine (Amph) on prepulse inhibition (PPI), a sensorimotor gating model in which the amplitude of the acoustic startle response (ASR) to a startling sound (pulse) is reduced when preceded immediately by a weaker stimulus (prepulse). Given that dopamine (DA) projections to the basolateral amygdala (BLA) are involved in the control of information processing, our aim was to investigate if intra-BLA administration of LNO would modify the disruption caused by the DA agonists, Amph, apomorphine (Apo) and quinpirole (QNP), on PPI. Male Wistar rats received bilateral intra-BLA microinjections (0.2 µL/min/side) of combined treatments (saline or LNO 11 µg followed by saline, QNP 3 µg, Apo 10 µg or Amph 30 µg). PPI was disrupted by intra-BLA Apo, QNP or Amph but not by LNO. Prior bilateral intra-BLA injection of LNO prevented the Apo- and QNP-induced disruption of PPI but did not affect that caused by Amph. APO- or QNP-induced increases in ASR to prepulse + pulse were also restored by LNO. Since local inhibition of NO formation affected the effects of direct, but not indirect, DA agonists, the results suggest that this modulation is not occurring at the level of DA release but may involve complex interactions with other neurotransmitter systems.

Advantages of a combined method of decalcification compared to EDTA

Decalcification of mineralized tissues is an essential step during tissue processing in the routine histopathology. The time required for complete decalcification, and the effect of decalcifier on cellular and tissue morphology are important parameters which influence the selection of decalcifying agents. In this study, we compared a decalcifying solution (ETDA) composed of both acid and chelating agents to a classical and well‐known decalcifying agent (EDTA). To this purpose, the optic density of bone radiographs, residual calcium analysis, bone sample weight, and histological and immunohistochemical analysis were performed. Our data suggest that, similarly to EDTA, the ETDA solution completely removes the calcium ions from the samples enabling easy sectioning. However, unlike the EDTA, this agent takes much less time. Furthermore, both agents showed comparable decalcification efficacy, and similarly, they did not produce cellular, tissue or antigenicity impairments. Therefore, ETDA may be a suitable option when it is necessary an association between a rapid and complete removal of calcium minerals, and a suitable preservation of structure and antigenicity of tissues. Microsc. Res. Tech. 78:111–118, 2015.

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.

Nanomedicine to Overcome Current Parkinson’s Treatment Liabilities: A Systematic Review

Nanoparticles might be produced and manipulated to present a large spectrum of properties. The physicochemical features of the engineered nanomaterials confer to them different features, including the ability to cross the blood–brain barrier. The main objective of this review is to present the state-of-art research in nano manipulation concerning Parkinson’s disease (PD). In the past few years, the association of drugs with nanoparticles solidly improved treatment outcomes. We systematically reviewed 28 studies, describing their potential contributions regarding the role of nanomedicine to increase the efficacy of known pharmacological strategies for PD treatment. Data from animal models resulted in the (i) improvement of pharmacological properties, (ii) more stable drug concentrations, (iii) longer half-live and (iv) attenuation of pharmacological adverse effects. As this approach is recent, with many of the described works being published less than 5 years ago, the expectancy is that this knowledge gives support to an improvement in the current clinical methods to the management of PD and other neurodegenerative diseases.

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.