Sheilla Da Silva Barroso
Sheilla Da Silva Barroso
Sheilla Da Silva Barroso
ARACAJU
JULHO – 2012
UNIVERSIDADE TIRADENTES
Orientadores
ARACAJU
JULHO – 2012
O AUTOR PERMITE A REPRODUÇÃO DE CÓPIAS OU PARTES DESTA
DISSERTAÇÃO DE MESTRADO SOMENTE PARA PROPÓSITOS ACADÊMICOS E
CIENTÍFICOS DESDE QUE A FONTE SEJA CITADA.
iii
EFEITOS COMPORTAMENTAIS DO EXERCÍCIO E DO EXTRATO DE
PRÓPOLIS VERMELHA SOBRE A DEGENERAÇÃO DE NEURÔNIOS
DOPAMINÉRGICOS EM RATOS
Aprovada por:
________________________________________________
Margarete Zanardo Gomes, D.Sc.
Orientadora
________________________________________________
Ricardo Luiz Cavalcanti de Albuquerque Júnior, D.Sc.
Orientador
_________________________________________________
Juliana Cordeiro Cardoso, D.Sc.
Examinadora
________________________________________________
Waldecy de Lucca Junior, D.Sc.
Examinador
________________________________________________
Katia Peres Gramacho D.Sc.
1º Suplente
________________________________________________
Barbara Lima Simioni Leite D.Sc.
2º Suplente
iv
“A melhor recompensa que a vida oferece é a
oportunidade de trabalhar duro em algo que valha a pena”
Theodore Roosevelt
v
AGRADECIMENTOS
A minha mãe pelo apoio, força, incentivo e amizade. Por estar ao meu lado me
encorajando nas horas difíceis e me aplaudindo nos momentos de glória. Dedico a
você este titulo, pois sem você nada disso seria possível. Amo muito você!
Aos meus orientadores Profª Dra. Margarete Zanardo Gomes e Prof. Dr. Ricardo
Luiz Cavalcanti de Albuquerque Júnior por acreditarem em mim e no futuro deste
projeto, por me mostrarem com sabedoria, discernimento, bom senso, dedicação o
caminho da ciência contribuindo para o meu crescimento profissional, sendo peças
fundamentais para a concretização desse momento especial. Neste agradecimento
tento expressar um pouco do carinho e respeito que tenho por vocês. Serei grata pelas
oportunidades aqui proporcionadas. Obrigada por transmitirem seus conhecimentos e
experiências profissionais, pelo acesso que me facilitou a uma pesquisa mais alargada
e pela sua crítica sempre tão enriquecedora.
A Camila, Tâssia, Tâmara, Rafaela, Nely, Fani, Jamile por estarem sempre ao meu
lado durante a realização do experimento, por me sempre socorrem com palavras
amigas, acalentadoras, pelo carinho, amizade que sempre me foi destinado e por
ajudar a tornar este sonho possível.
E por fim, muito obrigada a todos aqueles que contribuíram de alguma forma para a
realização deste trabalho.
vi
SUMÁRIO
INTRODUÇÃO ................................................................................................. 15
CAPITULO I ..................................................................................................... 17
1.3 Etiologia...................................................................................................... 21
CAPITULO II .................................................................................................... 54
RESUMO.......................................................................................................... 56
ABSTRACT ...................................................................................................... 57
INTRODUÇÃO ................................................................................................. 58
MÉTODOS ....................................................................................................... 59
RESULTADOS ................................................................................................. 62
DISCUSSÃO .................................................................................................... 64
CONCLUSÕES ................................................................................................ 66
REFERÊNCIAS ................................................................................................ 66
vii
CAPITULO III ................................................................................................... 71
ABSTRACT ...................................................................................................... 73
INTRODUCTION .............................................................................................. 74
RESULTS ......................................................................................................... 79
DISCUSSION ................................................................................................... 82
CONCLUSION ................................................................................................. 85
BIBLIOGRAPHY ............................................................................................... 86
ANEXOS .......................................................................................................... 94
viii
LISTA DE SIGLAS E ABREVIAÇÕES
6-OHDA : 6-hidroxidopamina
C: córtex
CAPE: Ácido fenil éster cafeico
COMT: Catecol-o-metiltransferase
DA: Neurônios dopaminérgicos
DAT: Transportador de dopamina
DMBA: 9,10-Dimetil-1,2-Benzantraceno
DP : Doença de Parkinson
EHPV: Extrato hidroetanólico de própolis vermelha
ERN’s: Espécies reativas de Nitrogênio
ERO’s: Espécies reativas de Oxigênio
GABA: Ácido Gama-aminobutirico
GDNF:Fator de Crescimento derivado da Glia
Glu: Neurônios Glutamatérgicos
GPe: Globo Pálido externo
GPi: Globo Pálido interno
L-DOPA: 3,4dihidroxifenilalanina
L-NOARG: NG-nitro-l-arginina
MAO-A: Monoamina Oxidase
MPTP: 1-metil-4-fenil-1,2,3,6-tetrahidropiridina
NAT: Transportador de noradrenalina
NB: Núcleos Basais
NO: óxido nítrico
NOS: Síntese de óxido nítrico
PPN: Núcleo Pendúculo Pontino
PQ: Paraquat
SN: Substância Negra
SNC: Sistema Nervoso Central
SNc: Substância Negra parte compacta
SNr: Substância Negra parte reticular
STN: Núcleo subtalâmico
STR: Estriado
Th: Tálamo
TH: Tirosina hidroxilase
VL: Núcleo Ventro-lateral
ix
LISTA DE TABELAS
CAPITULO II Pg.
CAPITULO III
Table 1: Percentage of TH immunoreactive (TH+) cells in the
substantia nigra (SN) from sham brains, 24 and 96 hours after 6-
OHDA injection into medial forebrain bundle. 6-OHDA induced an 80
ipsilateral decrease in the TH+ cells in a time-dependent and lateral
to medial manner. The 6-OHDA/L-NOARG treatment (100mg/kg
i.p.) retards the initial phases of the DA loss. * indicates difference
from control and # indicates difference from all other groups.
x
LISTA DE FIGURAS
CAPÍTULO I Pg.
CAPÍTULO II
CAPÍTULO III
xi
EFEITOS COMPORTAMENTAIS DO EXERCÍCIO E DO EXTRATO
DE PRÓPOLIS VERMELHA SOBRE A DEGENERAÇÃO DE
NEURÔNIOS DOPAMINÉRGICOS EM RATOS.
xii
BEHAVIORAL EFFECTS OF EXERCISE AND PROPOLIS
EXTRACT ON THE DEGENERATION OF DOPAMINERGIC
NEURONS IN RATS.
xiii
INTRODUÇÃO
INTRODUÇÃO
Além disso, o exercício físico tem sido proposto para amenizar os efeitos
comportamentais e neuroquímicos induzidos pela toxina monoaminérgica 6-
15
hidroxidopamina (6-OHDA) (TILLERSON et al., 2003; MABANDLA et al., 2004;
YOON et al., 2007; MABANDLA et al., 2009; TAJIRI et al., 2010; DUTRA et al.,
2012). Foi demonstrado que o exercício físico aumenta a expressão de fatores
neurotróficos de células gliais (GDNF) (COHEN et al., 2003), apresenta efeito
neuroprotetor (SMITH e ZIGMOND, 2003), contribui para ativação do sistema
dopaminérgico, aumenta a disponibilidade de dopamina no estriado e pode
reverter os déficits comportamentais relacionados ao movimento, ao equilíbrio
e ao desempenho da marcha (HATTORI e NAOI, 1994).
REVISÃO BIBLIOGRÁFICA
1 REVISÃO BIBLIOGRÁFICA
1.2 Fisiopatologia da DP
20
Figura 1. Modelo simplificado das vias direta e indireta dos núcleos da base. Em a) o
funcionamento em indivíduos normais, em b) o funcionamento em indivíduos com DP. Setas
pretas indicam conexões inibitórias (Gabaérgicas); setas brancas indicam conexões
excitatórias (Glutamatergicas). A mudança na espessura das setas indica a intensidade de
sinal transmitido pela sinapse, mostrando o desequilíbrio provocado pela falta de dopamina.
Abreviações: Substância negra compacta (SNc) e reticulada (SNr); Globo pálido externo (GPe)
e interno (GPi), núcleo subtalâmico (STN), via direta (D1), Via indireta (D2). Fonte: Adaptado de
GALVAN e WICHMANN 2008.
1.3 Etiologia
1.4 Tratamento
24
Conforme Shapira et al., (2009) o L-dopa pode ter um efeito tóxico em
modelos in vitro de cultura de células. Em contraste com os dados in vitro, há
pouca ou nenhuma evidência de toxicidade para os neurônios de DA
provocada pela L-dopa em modelos in vivo (MYTILINEOU et al., 2003). Por
exemplo, a administração de grandes quantidades de L-dopa em animais não
resultou em dano oxidativo ou degeneração das células da substância negra
(FERRARIO et al., 2004) e alguns têm mostrado efeito no fator de crescimento
(MENA et al., 1997).
26
Mesmo considerando os resultados favoráveis, clínicos e pré-clínicos,
pela utilização do exercício físico na DP, fazem-se necessários estudos que
determinem, por exemplo, o tipo de exercício físico, se aeróbio ou de
anaeróbio, o período para início da prática e a eficácia da associação com
outras estratégias terapêuticas.
O termo própolis é de origem grega pro (em defesa de), polis (cidade), o
que significa: “em defesa da cidade ou da colméia” (BANKOVA, 2000). Na
colônia as abelhas utilizam a própolis para esterilização, impermeabilização,
isolamento térmico, vedação e tratamento anti-séptico evitando assim, a
contaminação por animais invasores (MARCUCI, 1996; MARCUCCI, 2001;
LONGHINI, 2007).
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Farmacognosia, 17 (1), p.85-93, 2007.
SUNVISSON, H.; LÖKK, J.; ERICSON, K.; WINBLAD, B.; EKMAN, SL.
Changes in motor performance in persons with Parkinson's disease after
exercise in a mountain area. Journal of Neuroscience Nursing, 29 (4), p.255-
260, 1997.
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disease model of rats. Brain Research, 1310 (15), p.200-207, 2010.
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Puerarin protects dopaminergic neurons against 6-hydroxydopamine
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53
CAPITULO II
DOPAMINÉRGICOS EM RATOS.
Sergipe.
MÉTODOS
Animais
A amostra foi composta por 55 ratos machos adultos Wistar com peso
corporal 250-400g. Os animais foram alojados em gaiolas com cama de
maravalha, em temperatura controlada de (~±22°C), regime de luz com ciclo
claro-escuro de 12 h e livre acesso à água e comida. Todos os procedimentos
estiveram de acordo com as diretrizes do Manual de Princípios no Cuidado e
Uso de Animais do American College of Sports Medicine e seguiu a Lei nº.
6638 de 8 de maio de 1979 e o Decreto nº. 24645 de 10 de julho de 1934, com
aprovação do Comitê de Ética no Uso Animal da Universidade Tiradentes
(CEUA/UNIT/SE), sob parecer consubstanciado n° 170610.
59
Procedimento cirúrgico
Grupos Experimentais
Tratamentos
Análise Estatística
Tabela 1: Médias de comportamentos exibidos ± erro padrão da média para o teste de campo
aberto em animais não lesionados.
Lesão/P10
50,00 Lesão/P10 25,00 Lesão/Exc
Média das explorações verticais
b b Lesão/Exc Lesão/P10/Exc
Lesão/P10/Exc b
40,00 20,00
b
30,00 15,00
a
20,00 10,00
a a
10,00 5,00
0,00 0,00
63
DISCUSSÃO
64
ao EHPV, aumentou o comportamento de exploração horizontal e vertical em
relação ao grupo 6-OHDA tratado com veículo, restaurando a atividade motora
após lesão. Estes dados são indicativos de um efeito protetor do exercício
físico sobre a degeneração dopaminérgica. Em consonância, Smith e
Zigmond23 demonstraram que o exercício forçado pode reduzir vulnerabilidade
dos neurônios dopaminérgicos submetidos à lesão por 6-OHDA, sugerindo que
a proteção seja em parte pelo aumento na disponibilidade do fator neurotrófico
de células gliais (GDNF).
Outros estudos também mostraram que o exercício físico pode atenuar
as alterações comportamentais e neurobiológicas em roedores submetidos à
lesão dopaminérgica5,6. Faherty et al.,24 demonstraram que o exercício
realizado na roda de corrida exerce um efeito protetor contra morte neuronal
celular na substância negra induzida pela aplicação do 1-metil-4-fenil-1,2,3,6-
Tetrahidropiridina (MPTP) em camundongos. O’Dell et al., 25 verificaram que o
desempenho motor de ratos lesionados com a 6-OHDA e exercitados foi
melhor do que os ratos sedentarios, sugerindo que o exercício voluntario pode
facilitar a recuperação da lesão parcial da via nigroestriatal. No entanto, não há
relatos dos efeitos de exercício de natação associados ao Parkinson
experimental.
Em animais lesionados, o tratamento com EHPV isolado não produziu
alterações significativas quando comparado ao veículo, ou modificações nos
efeitos do exercício. Estudos anteriores mostraram que componentes bioativos
da própolis têm efeito neuroprotetor sobre a lesão de neurônios
dopaminérgicos. O ácido fenil cafeico (CAPE), por exemplo, apresentou efeito
neuroprotetor em baixas concentrações ante a toxina 6-OHDA em estudo
realizado com cultura de células26, além de prevenir a neurotoxidade causada
por glutamato27. Corroborando com outros estudos, Kurauchi et al.,28
descreveram que as ações protetoras do CAPE sobre a degeneração
dopaminérgica são mediadas tanto por heme-oxigenasase (HO-1) e fator de
crescimento da glia (BDNF), promovendo uma redução na síntese de óxido
nítrico e da expressão caspase 1 em estudos in vivo. Em um estudo recente foi
demonstrado que o CAPE diminuiu a neurodegeneração causada pelo MPTP
em camundongos29.
65
A não obtenção de modificações significativas no quadro após lesão
pela utilização do EHPV pode estar relacionada ao fato de a composição
química diferencial da própolis vermelha influenciar sua atividade biológica.
Esta variedade denominada de “própolis vermelha” foi classificada como o 13°
tipo de própolis Brasileira, por possuir uma coloração vermelha intensa e sua
composição química diferir dos 12 tipos de própolis brasileira apresentada por
Park et al.,30. Não obstante, o teste de campo aberto por si não permite afirmar
a não ocorrência de neuroproteção, sendo necessários mais estudos para a
compreensão da ação do EHPV no modelo de lesão pela aplicação de 6-
OHDA.
CONCLUSÕES
REFERÊNCIAS
2010; 1310:200-207.
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10. Pothakos K, Kurz MJ, Lau YS. Restorative effect of endurance exercise on
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x:xx-xx.
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22. Li XH, Wang JY, Gao G, Chang JY, Woodward DJ, Luo F. High-frequency
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25. O’Dell SJ, Gross NB, Fricks AN, Casiano BD, Nguyen TB, Marshall BJF.
69
29. Fontanilla CV, Ma Z, Wei X, Klotsche J, Zhao L, Wisniowski P, et al. Caffeic
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70
CAPITULO III
1, 2
Gomes, M. Z.; 1Barroso, S.S.; 2Del Bel, E.
1
Programa de Pós-Graduação em Saúde e Ambiente-Universidade Tiradentes,
SE, Avenida Murilo Dantas, 300, CEP 49032-490, SE, Brazil; 2Department of
MEF-Physiology, School of Odontology, Campus USP, Ribeirao Preto, SP
14040-904, Brazil.
E-mail: [email protected]
ABSTRACT
There is evidence that nitric oxide plays a role in the neurotransmitter balance
within the basal ganglia and in the pathology of Parkinson's disease. In the
present work we investigated the effects of a nitric oxide synthase (NOS)
inhibitor, NG-nitro-l-arginine (L-NOARG), on both the dopaminergic neuronal
loss and the neuronal NOS cell density after medial forebrain bundle 6-
hydroxydopamine (6-OHDA) lesion, as a function of the time. We analyzed the
dopaminergic (DA) neuronal loss through tyrosine hydroxylase
immunohistochemistry (TH), 24 and 96 hours and also 15 and 35 days after 6-
OHDA or 6-OHDA/L-NOARG. The nitrergic system was evaluated using an
antibody against the neuronal NOS isoform. 6-OHDA induced a striatal increase
in the density of neuronal NOS from 24 hours until 35 days after lesion that
inversely correlated with progressive DA cell loss in the susbtantia nigra (SN),
lateral to medial regions. Acute treatment with the L-NOARG significantly
reduced 6-OHDA effects on TH and NOS. The results suggest that nitric oxide
synthase inhibition may retards the toxic effects of 6-OHDA on dopaminergic
cell bodies in SN and on neuronal nitric oxide synthase cell density in the rat
brain.
Animals
Drugs
Surgical procedure
75
Nigrostriatal lesions were produced by microinjecting 6-OHDA with 0.02%
ascorbic acid (4 μl, 8 μg/μl, RBI-Sigma) in 0.9% saline into the right medial
forebrain bundle with a 50 μl Hamilton syringe.
The stereotaxic coordinates, according to the Paxinos; Watson’s (1998)
rat brain atlas, were: anterior: −4.4; lateral: −1.2, and vertical: −8.2 from
bregma. The microinjections were performed in a rate of 1 µl/min with an
infusion pump (Scientific, USA), and the needle was left in place for an
additional 180s to prevent reflux. The movement of an air bubble inside the PE-
10 polyethylene tubing connecting the micro-syringe with the needle confirmed
drug flow. Sham-operated control animals were submitted to the same
procedure but were injected with 4 μl of vehicle instead of the neurotoxin.
Experimental groups
Histological analysis
77
Sections were mounted onto gelatin-coated glass slides, dehydrated in
ethanol, cleared in xylene, and cover-slipped for microscopic observations.
Immunopositive cells were revealed by a brown reaction product. Tissue from
sham and lesion groups was always processed at the same time. In all
experiments tissues from every group were always processed in the same
assay.
Quantitative morphology
78
calculated from results obtained in each brain side. Results of the TH+ are
expressed as a percentage of the same side in sham brains.
Statistical analysis
RESULTS
Rotational behavior
Quantitative morphology
79
Loss of TH+ appears first in the lateral SN (26.1% ± 10 remaining cells)
24 hours after 6-OHDA (table 1), suffering a further decrease with 96 hours
(absence of cells). Still 96 hours after surgery, it was also observed a decrease
in the TH+ at ventral (9.9% ± 4 remaining) and dorsal SN (41.2% ± 7).
L-NOARG treatment attenuated TH+ loss in the lateral SN 24 hours
(69.6% ± 8.6 remaining, P= 0.03; F= 3.8) and in the ventral SN 96 hours after 6-
OHDA (39.7% ± 11.4, P<0.0001; F=32.4, one way ANOVA followed by Duncan
post test). However, this 6-OHDA/L-NOARG treatment did not modified TH+ in
the dorsal SN 96 hours after 6-OHDA (33.4% ± 6.7 remaining, P=0.02; F= 4.2).
Table 1: Percentage of TH immunoreactive (TH+) cells in the substantia nigra (SN) from sham
brains, 24 and 96 hours after 6-OHDA injection into medial forebrain bundle. 6-OHDA induced
an ipsilateral decrease in the TH+ cells in a time-dependent and lateral to medial manner. The
6-OHDA/L-NOARG treatment (100mg/kg i.p.) retards the initial phases of the DA loss. *
indicates difference from control and # indicates difference from all other groups.
6-OHDA 6-OHDA/L-NOARG
Lateral SN 26.1%±9.7# 69.6%±8.6
24 hours Medial SN 72.6%±10.1 75.3%±13.6
Dorsal SN 69.6%±17.4 75.5%±12.2
Lateral SN 0.0±0.0 0.0±0.0
96 hours Medial SN 9.9%±5.6# 39.7%±11.4
Dorsal SN 41.2%±12.0* 33.4%±6.7*
Density of striatal TH+ fibers was unchanged both 24 and 96 hours after
6-OHDA (caudate/putamen and acumbens, results not showed). We also
evaluated TH+ 20 and 35 days after neurotoxin injection. At this point it was
observed massive loss, with lacking of stained cells and terminals ipsilateral to
lesion in both groups 6-OHDA and 6-OHDA/L-NOARG.
Either saline or L-NOARG treatment (i.p) associated with saline
microinjection (sham) did not elicit difference in TH+ cells or terminal density in
any measured region (results not presented). Also, contra-lateral side of TH
stained sections did not show significant differences when compared with the
same side in sham rats.
80
Time course of nNOS (+) modifications after 6-OHDA and the effects of the
L-NOARG treatment
nNOS+ was detected in the basal ganglia, including the SN and striatum.
Quantitative analysis revealed that L-NOARG per se (saline/L-NOARG) did not
alter the density of positive neurons. As the nNOS+ neurons could be finding
only in the dorsal tier of SN, we counted the number of nNOS+ neurons no
more than in this sub region and in the dorsal striatum.
In the SN, nNOS+ cell counts were unchanged 24 and 96 hours after 6-
OHDA or 6-OHDA/L-NOARG but decreased 20 and 35 days after toxin
injection in both groups (P< 0.0001, F= 42 and P< 0.0001, F= 39, respectively,
one way ANOVA followed by Duncan post test).
On the other hand, 6-OHDA induced an increase in the density of
nNOS(+) cells in the striatum ipsi and contralateral 24 hours to 35 days after
neurotoxin injection when compared with sham rats (figure 1). 6-OHDA/L-
NOARG treatment prevents nNOS+ increase in both sides 24 hours (P= 0.002,
F= 44.6 and P=0.01, F= 4.3) and 35 days (P= 0.03, F= 4.8 and P= 0.002, F=
44.6, figure 1) after lesion.
81
Figure 1: Mean density of nNOS+ cells in the striatum of rats sacrificed 24 hours and 35 days
after injection of 6-OHDA or saline into medial forebrain bundle. White columns represent the
mean density of nNOS+ cells in the sham rats, black columns represent 6-OHDA group and
dashed columns represents 6-OHDA/L-NOARG group. Upper panel shows the ipsilateral
counts to neurotoxin injection, while lower panel illustrates contralateral results. Rats received
one single application of L-NOARG 100mg/kg or saline i.p. after surgery. * indicates significant
difference (P < 0.05, MANOVA followed by One way ANOVA, Duncan’s post test).
DISCUSSION
The DA cell loss in the SN as a function of the time has been well
described in the 6-OHDA-induced medial forebrain bundle lesion. First signs of
neuronal death appear since 24 hours after toxin injection while the peak of
82
neurotoxicity occurs with 48 hours (ZUCH et al., 2000). At this time, we
observed a lateral – to medial gradient of TH+ loss in the SN after 6-OHDA from
24 to 96 hours. Similar results were described in PD (HU et al. 2006;
HUTCHINSON et al. 1999, 2003) and in an early rat 6-OHDA model (GOMES
et al., 2008, DEBEIR et al., 2005; OLDS et al., 2006; FALLON; MOORE, 1978).
Cytotoxicity of 6-OHDA is due to its pro-oxidant activity; the toxin
undergoes rapid auto-oxidation in the extracellular space, thus promoting a high
rate of reactive oxygen species formation (HANROTT et al., 2006), which is
associated with activation of the apoptotic cascade (BERNSTEIN et al., 2011;
FUJITA; NAKABEPPU; NODA, 2011). The direct injection of 6-OHDA into the
SNc and/or MFB, yields a virtually immediate lesion of dopaminergic cell
bodies, followed by degeneration of striatal terminals (MEREDITH, 2008).
L-NOARG treatment significantly decreases the TH+ cell loss. Previous
studies have shown that NOS inhibition can protect DA cells against MPTP and
6-OHDA-induced neurotoxicity in experimental animals (GOMES et al., 2008;
SCHULZ et al. 1995; HANTRAYE et al. 1996, WATANABE et al., 2007). This
protective effect of NOS inhibition by L-NOARG can be due to activity of NO on
dopamine transporter (DAT). Nitric oxide produced from L-arginine increases
DAT activity and so that the 6-OHDA access (VOLZ; SCHENK 2004).
Alternatively, L-NOARG can inhibit biochemical interaction of NO with the 6-
OHDA that leads to cellular oxidation (SMITH; CASS, 2006; HENZE et al. 2005;
HANROTT et al. 2006; ANTUNES et al. 2005). On the other hand, TH (+) in the
SN was absent after 15 days in all 6-OHDA lesioned groups, indicating that a
single application of L-NOARG (100mg/kg) could not stop neurotoxicity.
TH (+) was not modified in the striatum 24 – 96 hours. DA depletion can
be observed in the striatum only after about 80% of neuronal cell death that
results in dennervation (DEUMENS et al., 2002). A compensatory post synaptic
hipersensitivity mechanism, by increasing in the number and affinity of
receptors, has been described after 15 days (for review see SCHWARTING;
HUSTON, 1996). As a result, a DA agonist administration, like apomorfine,
induces rotational behavior. In agreement with SN and striatal results (missing
of TH+ in the 6-OHDA and 6-OHDA/L-ANORG groups 15 days after surgery),
rotational behavior was not modified by L-NOARG.
83
Effects of the L-NOARG treatment on time-course of nNOS (+)
modifications after 6-OHDA
84
2002; WEST; GALLOWAY 1998) and the increased nNOS cell density could
result from a compensatory augmentation in the activity of striatal nNOS+
interneurons, which are excited by an intact nigrostriatal pathway through a D1-
like receptor mechanism (CENTONZE et al., 2002). In agreement, Salin et al
(2009) have described reorganization between striatal interneurons (nNOS
positive neurons) and projection neurons after 6-OHDA. It supports the notion of
an inhibitory modulatory role exerted by endogenous NO on control striatal
projection cells. Still, there are indications in the literature of an increase in the
nNOS cell density or NADPH-diaphorase activity in the striatum after chronic
deafferentation (GOMES et al., 2008; SANCESARIO et al., 2004; MORTON et
al., 1993; GOMES; DEL BEL, 2003, MURAMATSU et al., 2002).
L-NOARG treatment reverses striatal 6-OHDA-induced nNOS+ increase,
24 hours to 35 days. Since L-NOARG per se did not affect nNOS+ (saline
injected rats), we can attribute this effect as another evidence of its protective
role under DA lesion. Galati et al. (2008) suggested a functional cross-talk
between NO spontaneously active interneurons and projection neurons, which
becomes critical in the parkinsonian state. Then, this neuromodulatory role of
NO could be permanent until 35 days after lesion.
CONCLUSION
85
Acknowledgements
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