Complement activation in the Parkinson's disease substantia nigra: an immunocytochemical study

BACKGROUND: Inflammatory processes are increased in the Parkinson's disease (PD) brain. The long-term use of nonsteroidal anti-inflammatory drugs has been associated, in retrospective studies, with decreased risk for PD, suggesting that inflammation may contribute to development of this disorder. The objective of this study was to determine the extent of complement activation, a major inflammatory mechanism, in PD. METHODS: Substantia nigra specimens from young normal subjects (n = 11–13), aged normal subjects (n = 24–28), and subjects with PD (n = 19–20), Alzheimer's disease (AD; n = 12–13), and dementia with Lewy bodies (DLB; n = 9) were stained for iC3b and C9, representing early- and late-stage complement activation, respectively. Numbers of iC3b(+), C9(+), and total melanized neurons in each section were counted in a blinded fashion. Nonparametric analyses were used to evaluate differences between groups and to evaluate correlations between complement staining, numbers of melanized neurons, and the duration of PD. RESULTS: Lewy bodies in both PD and DLB specimens stained for iC3b and C9. Staining was also prominent on melanized neurons. The percentage of iC3b(+ )neurons was significantly increased in PD vs. aged normal and AD specimens, and in young normal vs. aged normal specimens. C9 immunoreactivity was significantly increased in PD vs. AD specimens, but unlike iC3b, the increased C9 staining in PD and young normal specimens did not achieve statistical significance vs. aged normal specimens. iC3b and C9 staining in PD specimens was not correlated with the numbers of remaining melanized neurons, nor with the duration of PD. CONCLUSION: Complement activation occurs on Lewy bodies and melanized neurons in the PD substantia nigra. Early complement activation (iC3b) is increased on melanized neurons in PD vs. aged normal specimens, and late-stage complement activation (C9) also tends to increase. This latter finding suggests that complement activation may contribute to loss of dopaminergic neurons in some individuals with PD. Complement activation on melanized neurons appears to decrease with normal aging, suggesting a possible neuroprotective role for this process in the normal substantia nigra

Dynamic Measurements of Cerebral Pentose Phosphate Pathway Activity In Vivo Using [1,6- 13 C 2 ,6,6- 2 H 2 ] Glucose and Microdialysis

Cerebral pentose phosphate pathway (PPP) activity has been linked to NADPH-dependent anabolic pathways, turnover of neurotransmitters, and protection from oxidative stress. Research on this potentially important pathway has been hampered, however, because measurement of regional cerebral PPP activity in vivo has not been possible. Our efforts to address this need focused on the use of a novel isotopically substituted glucose molecule, [1,6- 13 C 2 ,6,6- 2 H 2 ]glucose, in conjunction with microdialysis techniques, to measure cerebral PPP activity in vivo, in freely moving rats. Metabolism of [1,6- 13 C 2 ,6,6- 2 H 2 ]glucose through glycolysis produces [3- 13 C]lactate and [3- 13 C,3,3- 2 H 2 ]lactate, whereas metabolism through the PPP produces [3- 13 C,3,3- 2 H 2 ]lactate and unlabeled lactate. The ratios of these lactate isotopomers can be quantified using gas chromatography/mass spectrometry (GC/MS) for calculation of PPP activity, which is reported as the percentage of glucose metabolized to lactate that passed through the PPP. Following addition of [1,6- 13 C 2 ,6,6- 2 H 2 ]glucose to the perfusate, labeled lactate was easily detectable in dialysate using GC/MS. Basal forebrain and intracerebral 9L glioma PPP values (mean ± SD) were 3.5 ± 0.4 (n = 4) and 6.2 ± 0.9% (n = 4), respectively. Furthermore, PPP activity could be stimulated in vivo by addition of phenazine methosulfate, an artificial electron acceptor for NADPH, to the perfusion stream. These results show that the activity of the PPP can now be measured dynamically and regionally in the brains of conscious animals in vivo.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65845/1/j.1471-4159.1995.64031336.x.pd

Plaque complement activation and cognitive loss in Alzheimer's disease

<p>Abstract</p> <p>Background</p> <p>Complement activation is increased in Alzheimer's disease (AD), but its significance is unclear. The objective of this study was to determine the relationship between complement activation and cognition during the development of AD.</p> <p>Methods</p> <p>iC3b, C9, Bielschowsky, and Gallyas staining was performed on aged normal (n = 17), mild cognitively impaired (n = 12), and AD (n = 17–18) inferior temporal gyrus specimens. Plaques were counted in 10× fields with high numbers of Bielschowsky-stained plaques. One-way ANOVA was used to determine between-group differences for plaque counts and measures of cognitive function, and linear regression was used to evaluate global cognition as a function of Bielschowsky-stained plaques. Terms for iC3b- and C9-stained plaques were then added sequentially as additional predictors in a "mediation analysis" model.</p> <p>Results</p> <p>Complement was detected on plaques in all groups, and on neurofibrillary tangles only in AD specimens. iC3b, C9, and Bielschowsky-stained plaque counts increased 2.5- to 3-fold in AD vs. other groups (all <it>p </it>≤ 0.01). C9 staining was present on some diffuse plaques, as well as on neuritic plaques. Bielschowsky-stained and complement-stained plaque counts were highly correlated, and were negatively correlated with cognitive measures. When the Bielschowsky plaque count was used as a predictor, its correlations with cognitive measures were statistically significant, but when iC3b and C9 plaque counts were added as additional predictors, these correlations were no longer significant. This loss of significance was attributed to multicollinearity, i.e., high correlations between Bielschowsky-stained and complement-stained plaque counts.</p> <p>Conclusion</p> <p>Both early-stage (iC3b) and late-stage (C9) complement activation occurs on neocortical plaques in subjects across the cognitive spectrum; contrary to previous reports, C9 is present on some diffuse plaques. Because of high correlations between complement-stained and Bielschowsky-stained plaque counts, quantitative assessment of the extent to which complement activation may mediate the relationship between plaques and cognitive function could not be performed. Additional studies with animal models of AD (if late-stage complement activation can be demonstrated), or possibly a trial in AD patients with an inhibitor of late-stage complement activation, may be necessary to determine the significance of this process in AD.</p

Long-lasting effects of escalating doses of d-amphetamine on brain monoamines, amphetamine-induced stereotyped behavior and spontaneous nocturnal locomotion

The repeated intermittent administration of relatively low doses of amphetamine (AMPH) produces an enduring hypersensitivity to the motor stimulant effects of AMPH (behavioral sensitization), and this accompanied by enhanced mesotelencephalic dopamine (DA) utilization/release. In contrast, chronic treatment with very high doses of AMPH does not produce sensitization, but is neurotoxic, resulting in the depletion of brain DA (and often other monoamines). However, gradually escalating doses of AMPH provide protection against the neurotoxic effects of higher doses given later. Therefore, the purpose of the present experiment was to determine if a regimen of gradually escalating doses of AMPH, culminating in much higher doses than usually used to study sensitization, would produce neural and behavioral changes associated with AMPH neurotoxicity (DA depletion) or behavioral sensitization (increased DA utilization). Female rats were given 60 injections (2/day) of increasing (1 to 10 mg/kg) doses of d-AMPH, culminating in rats receiving 20 mg/kg/day for four consecutive days. This treatment did not deplete brain DA or serotonin, but did produce a long-lasting enhancement (at least 12 days) in striatal and nucleus accumbens DOPAC concentrations, and DOPAC/DA ratios. These neurochemical changes were accompanied by an enduring hypersensitivity to the stereotypy-producing effects of a subsequent AMPH `challenge.' In contrast to this enhanced response to a challenge, AMPH-pretreated rats showed a marked reduction in spontaneous nocturnal motor activity. It is concluded that rats can be given escalating doses of AMPH, which mimic to some extent the AMPH `runs' common in addicts and that this produces neural and behavioral changes consistent with the development of sensitization; not neurotoxicity.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26763/1/0000315.pd

The effects of four days of continuous striatal microdialysis on indices of dopamine and serotonin neurotransmission in rats

The effects of 4 days of continuous microdialysis with a small-diameter concentric-style probe on indices of striatal dopamine (DA) and serotonin neurotransmission were assessed. It was found that over 4 days of dialysis, there was a marked time-dependent decrease in the basal concentrations of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in dialysate and in amphetamine-stimulated DA release. In contrast, there was no decrease in basal DA or in the ability of cocaine to elevate the concentration of DA in dialysate over the same period of time. There were only very modest changes in dialysate levels of the serotonin metabolite, 5-hydroxyindoleacetic acid (5-HIAA), relative to the marked changes in DA metabolites. It is suggested that 4 days of continuous dialysis does not result in a non-specific decrease in diffusibility of these compounds into the dialysis probe, but that the changes are more likely due to probe-induced damage to the nigrostriatal DA system. It is also suggested that a "stable" basal concentration of DA in dialysate is an especially poor indicator of the integrity of the dopaminergic input to the striatum. The implications of these findings for within-subjects design microdialysis experiments are discussed.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/28999/1/0000028.pd

Susceptibility to sensitization. I. Sex differences in the enduring effects of chronic -amphetamine treatment on locomotion, stereotyped behavior and brain monoamines

There are sex differences in a number of behaviors elicited by amphetamine (AMPH). The purpose of the present experiment was to determine if there are also sex differences in the sensitization of the locomotor activity and stereotypy produced by repeated intermittent AMPH treatment, and whether this is accompanied by sex differences in dopamine (DA) metabolism. It was found that female rats showed greater and more rapid sensitization of locomotor activity and stereotyped behavior than males. In addition, prior exposure to AMPH was associated with an elevation in resting striatal dihydroxyphenylacetic acid (DOPAC) to DA ratios in female, but not male rats, suggesting a sex difference in one neurochemical correlate of sensitization. As a group, males were more variable and heterogeneous in their response to repeated AMPH treatment, because they were divisible into two neurochemically distinct subgroups on the basis of their change in behavior and females were not. This heterogeneity may make it more difficult to identify neurochemical correlates of sensitization in males. It is suggested that there is a sex difference in the responsiveness of brain DA systems to repetitive activation, and this contributes to individual variation in the susceptibility to sensitization.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/27142/1/0000136.pd

Susceptibility to sensitization. II. The influence of gonadal hormones on enduring changes in brain monoamines and behavior produced by the repeated administration of -amphetamine or restraint stress

Repeated amphetamine use produces an enduring sensitization of brain dopamine (DA) systems and behavior. Repeated exposure to stress can also produce sensitization, and amphetamine and stress may be interchangeable in this regard. There is, however, enormous individual variation in the susceptibility to sensitization by either stimulants or stress. The purpose of the present study was to determine if endogenous gonadal hormones contribute to individual variation in the sensitization of stereotyped behaviors, locomotion or regional brain monoamine metabolism. It was found that removal of testicular hormones by castration of male rats facilitated the behavioral sensitization produced by either repeated amphetamine treatment or repeated restraint stress, but ovariectomy of female rats was without effect. Prior exposure to amphetamine enhanced striatal homovanillic acid (HVA) levels and dihydroxyphenylacetic acid to DA and HVA to DA ratios in intact female, ovariectomized female and castrated male rats, but not gonadally-intact male rats. As a group, intact males were particularly heterogeneous because they were divisible into two neurochemically distinct subgroups based on their degree of behavioral sensitization, and the other groups were not. It is suggested that individual differences in the sensitization of brain DA systems and behavior produced by repeated exposure to amphetamine or stress may be due in part to individual differences in the concentration of a testicular hormone.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/27143/1/0000137.pd

Basal extracellular dopamine in the nucleus accumbens during amphetamine withdrawal: a `no net flux' microdialysis study

The basal extracellular concentration of dopamine in the nucleus accumbens was quantified using the `no net flux' microdialysis method, in rats undergoing withdrawal from -amphetamine. Rats were initially pretreated with saline, or an escalating dose amphetamine regimen known to produce a robust withdrawal syndrome, and extracellular dopamine was quantified 3 or 28 days after the last pretreatment injection. There was no effect of amphetamine pretreatment on the basal extracellular concentration of dopamine in the nucleus accumbens, or on the `in vivo recovery' of dopamine, estimated by `no net flux' microdialysis. It is suggested that amphetamine withdrawal is not necessarily accompanied by changes in the basal extracellular concentration of dopamine in the nucleus accumbens.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/30384/1/0000002.pd

Cellular immune response to intrastriatally implanted allogeneic bone marrow stromal cells in a rat model of Parkinson's disease

<p>Abstract</p> <p>Background</p> <p>Marrow stromal cells (MSC), the non-hematopoietic precursor cells in bone marrow, are being investigated for therapeutic potential in CNS disorders. Although <it>in vitro </it>studies have suggested that MSC may be immunologically inert, their immunogenicity following transplantation into allogeneic recipients is unclear. The primary objective of this study was to investigate the cellular immune response to MSC injected into the striatum of allogeneic recipients (6-hydroxydopamine [6-OHDA]-hemilesioned rats, an animal model of Parkinson's disease [PD]), and the secondary objective was to determine the ability of these cells to prevent nigrostriatal dopamine depletion and associated motor deficits in these animals.</p> <p>Methods</p> <p>5-Bromo-2-deoxyuridine (BrdU) – labeled MSC from two allogeneic sources (Wistar and ACI rats) were implanted into the striatum of adult Wistar rats at the same time as 6-OHDA was administered into the substantia nigra. Behavioral tests were administered one to two weeks before and 16–20 days after 6-OHDA lesioning and MSC transplantation. Immunocytochemical staining for T helper and T cytotoxic lymphocytes, microglia/macrophages, and major histocompatibility class I and II antigens was performed on post-transplantation days 22–24. MSC were detected with an anti-BrdU antibody.</p> <p>Results</p> <p>Tissue injury due to the transplantation procedure produced a localized cellular immune response. Unexpectedly, both sources of allogeneic MSC generated robust cellular immune responses in the host striatum; the extent of this response was similar in the two allograft systems. Despite these immune responses, BrdU⁺cells (presumptive MSC) remained in the striatum of all animals that received MSC. The numbers of remaining MSC tended to be increased (<it>p </it>= 0.055) in rats receiving Wistar MSC versus those receiving ACI MSC. MSC administration did not prevent behavioral deficits or dopamine depletion in the 6-OHDA-lesioned animals.</p> <p>Conclusion</p> <p>MSC, when implanted into the striatum of allogeneic animals, provoke a marked immune response which is not sufficient to clear these cells by 22–24 days post-transplantation. In the experimental paradigm in this study, MSC did not prevent nigrostriatal dopamine depletion and its associated behavioral deficits. Additional studies are indicated to clarify the effects of this immune response on MSC survival and function before initiating trials with these cells in patients with PD or other neurodegenerative disorders.</p

A microdialysis study of ventral striatal dopamine during sexual behavior in female Syrian hamsters

Microdialysis was used to study the effects of exposure to a male hamster on extracellular concentrations of dopamine, dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 5-hydroxyindole acetic acid (5-HIAA) in the ventral striatum of ovariectomized female Syrian hamsters pretreated with either estradiol and progesterone, or a similar regimen of oil injections. The hormone-treated females showed high levels of lordosis throughout the hour of exposure to the male. In hormone-treated females, there was a rapid elevation of dialysate dopamine within the first 15 min of exposure to the male. Dialysate dopamine gradually declined over the next 45 min, though remaining significantly above baseline during the entire period of exposure to the male. None of the oil-treated females showed any indication of lordosis, and the addition of the male produced only a small increase in dopamine at 30 min, after which dopamine returned to pre-male basal levels. DOPAC, HVA, and 5-HIAA were all elevated following introduction of the male for both groups of females. These results suggest that ovarian hormones modulate the responsivity of ventral striatal dopamine to incentive stimuli associated with mating behavior in females, although extracellular levels of dopamine in the ventral striatum do not seem to be directly coupled to the display of lordosis.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/30731/1/0000380.pd