Placental Transfusion, Timing of Plastic Wrap or Bag Placement, and Preterm Neonates.

OBJECTIVE: Compare delivery room practices and outcomes of infants born at less than 32 weeks' gestation or less than 1,500 g who have plastic wrap/bag placement simultaneously during placental transfusion to those receiving plastic wrap/bag placement sequentially following placental transfusion. STUDY DESIGN: Retrospective analysis of data from a multisite quality improvement initiative to refine stabilization procedures pertaining to placental transfusion and thermoregulation using a plastic wrap/bag. Delivery room practices and outcome data in 590 total cases receiving placental transfusion were controlled for propensity score matching and hospital of birth. RESULTS: The simultaneous and sequential groups were similar in demographic and most outcome metrics. The simultaneous group had longer duration of delayed cord clamping compared with the sequential group (42.3 ± 14.8 vs. 34.1 ± 10.3 seconds, p < 0.001), and fewer number of times cord milking was performed (0.41 ± 1.26 vs. 0.86 ± 1.92 seconds, p < 0.001). The time to initiate respiratory support was also significantly shorter in the simultaneous group (97.2 ± 100.6 vs. 125.2 ± 177.6 seconds, p = 0.02). The combined outcome of death or necrotizing enterocolitis in the simultaneous group was more frequent than in the sequential group (15.3 vs. 9.3%, p = 0.038); all other outcomes measured were similar. CONCLUSION: Timing of plastic wrap/bag placement during placental transfusion did affect duration of delayed cord clamping, number of times cord milking was performed, and time to initiate respiratory support in the delivery room but did not alter birth hospital outcomes or respiratory care practices other than the combined outcome of death or necrotizing enterocolitis. KEY POINTS: · Plastic bag placement during placental transfusion is effective in stabilization of preterms.. · Plastic bag placement after placental transfusion is effective in stabilization of preterms.. · Plastic bag placement during placental transfusion and risk of death or necrotizing enterocolitis needs additional study..

A gene-based recessive diplotype exome scan discovers FGF6, a novel hepcidin-regulating iron-metabolism gene.

Standard analyses applied to genome-wide association data are well designed to detect additive effects of moderate strength. However, the power for standard genome-wide association study (GWAS) analyses to identify effects from recessive diplotypes is not typically high. We proposed and conducted a gene-based compound heterozygosity test to reveal additional genes underlying complex diseases. With this approach applied to iron overload, a strong association signal was identified between the fibroblast growth factor-encoding gene, FGF6, and hemochromatosis in the central Wisconsin population. Functional validation showed that fibroblast growth factor 6 protein (FGF-6) regulates iron homeostasis and induces transcriptional regulation of hepcidin. Moreover, specific identified FGF6 variants differentially impact iron metabolism. In addition, FGF6 downregulation correlated with iron-metabolism dysfunction in systemic sclerosis and cancer cells. Using the recessive diplotype approach revealed a novel susceptibility hemochromatosis gene and has extended our understanding of the mechanisms involved in iron metabolism.

Alpha-Synuclein contributes to GSK-3beta-catalyzed Tau phosphorylation in Parkinson's disease models.

We have shown in the parkinsonism-inducing neurotoxin MPP(+)/MPTP model that alpha-Synuclein (alpha-Syn), a presynaptic protein causal in Parkinson's disease (PD), contributes to hyperphosphorylation of Tau (p-Tau), a protein normally linked to tauopathies, such as Alzheimer's disease (AD). Here, we investigated the kinase involved and show that the Tau-specific kinase, glycogen synthase kinase 3beta (GSK-3beta), is robustly activated in various MPP(+)/MPTP models of Parkinsonism (SH-SY5Y cotransfected cells, mesencephalic neurons, transgenic mice overexpressing alpha-Syn, and postmortem striatum of PD patients). The activation of GSK-3beta was absolutely dependent on the presence of alpha-Syn, as indexed by the absence of p-GSK-3beta in cells lacking alpha-Syn and in alpha-Syn KO mice. MPP(+) treatment induced translocation and accumulation of p-GSK-3beta in nuclei of SH-SY5Y cells and mesencephalic neurons. Through coimmunoprecipitation (co-IP), we found that alpha-Syn, pSer396/404-Tau, and p-GSK-3beta exist as a heterotrimeric complex in SH-SY5Y cells. GSK-3beta inhibitors (lithium and TDZD-8) protected against MPP(+)-induced events in SH-SY5Y cells, preventing cell death and p-GSK-3beta formation, by reversing increases in alpha-Syn accumulation and p-Tau formation. These data unveil a previously unappreciated role of alpha-Syn in the induction of p-GSK-3beta, and demonstrate the importance of this kinase in the genesis and maintenance of neurodegenerative changes associated with PD.

Economic Evaluation and Systematic Review of Publicly Available Workers' Compensation Practice Details and Mod Rate Calculators Applied to Upper Midwest Agriculture.

Background: Agricultural employment is one of the most dangerous occupations in the United States. Workers' compensation coverage requirements for agricultural work vary from state to state, and experience modifier rates (E-mods) affecting insurance premiums sometimes vary drastically across state lines and according to claim severities and farm sizes. We proposed to develop an interactive software application that would educate farmers on the impact of employee time loss on annual E-mod factor change specific to their geographic location and farm size.Methods: We conducted a comparative analysis of workers' compensation formulations, including E-mods among Upper Midwestern states. We performed sensitivity analysis of the formulas to claim amount and payroll to highlight differences related to claim severity and to farm size.Results: The state to state variation and remarkable complexity of these formulas was confirmed. E-Mod factors are shown to increase substantially across states with both claim size and payroll, though are found to be similar across Wisconsin and Minnesota which were examined in detail.Conclusions: The findings confirm that creating a nationally applicable interactive educational software tool for farmers and ranchers to view hypothetical rate changes by inputting on-farm injury scenarios represents a significant challenge and that educational outreach coupled with the use of commercial software, especially as less costly options become available, may serve the role of minimizing misunderstandings by current producers as may other informational sources.

c-Fos expression associated with reinstatement of cocaine-seeking behavior by response-contingent conditioned cues.

The capability of cocaine cues to generate craving in cocaine-dependent humans, even after extended abstinence, is modeled in rats using cue reinstatement of extinguished cocaine-seeking behavior. We investigated neural activity associated with incentive motivational effects of cocaine cues using c-fos mRNA and Fos protein expression as markers. Unlike preceding studies, we used response-contingent presentation of discrete cues to elicit cocaine seeking. Rats were first trained to press a lever, resulting in cocaine reinforcement and light and tone cues. Rats then underwent extinction training, during which lever presses decreased. On the test day, rats either received response-contingent cocaine cues or received no cues. The cues reinstated extinguished cocaine-seeking behavior on the test day. In general, cue-elicited c-fos mRNA and protein expression were similar and both were enhanced in the prefrontal cortex, ventral tegmental area (VTA), dorsal striatum, and nucleus accumbens. Cues elicited more widespread Fos protein expression relative to our previous research in which cues were presented noncontingently without prior extinction training, including increases in the VTA, substantia nigra, ventral subiculum, and lateral entorhinal cortex. We also observed a correlation between cocaine-seeking behavior and Fos in the agranular insula (AgI) and basolateral amygdala (BLA). The findings suggest that connections between BLA and AgI play a role in cue-elicited incentive motivation for cocaine and that reinstatement of cocaine seeking by response-contingent cues activates a similar corticolimbic circuit as that observed with other modes of cue presentation; however, activation of midbrain and ventral hippocampal regions may be unique to reinstatement by response-contingent cues.

Dopamine D3 receptor agonists for protection and repair in Parkinson's disease.

Parkinson's disease is a severe, age-related neurodegenerative disorder in which a loss of substantia nigra-derived dopaminergic pathways to the striatum triggers profound motor perturbation, as well as cognitive, sensory and mood deficits. Although the dopamine precursor, L-dopa, is effective in the short-term in relieving motor dysfunction, it does not stop the progressive disappearance of dopaminergic neurons, encouraging interest in alternative therapeutic strategies. Dopaminergic agonists, such as pramipexole, appear to have neuroprotective and neurorestorative actions based on clinical and, most convincingly, experimental work. The role of specific dopaminergic receptor subtypes is an important issue, especially with respect to new drug development. Of particular interest, dopamine D3 receptors contribute to the beneficial influence of dopaminergic agonists for the protection and restoration of dopaminergic pathways in Parkinson's disease.

Dopamine D3 receptor antagonists as therapeutic agents.

The behavioral and pathophysiological role of the dopamine D(3) receptor, which was deduced from anatomical, lesion and drug treatment studies in the ten years following cloning of the receptor, indicated that its functions differed from those of the D(2) receptor. There is increasingly strong evidence that D(3) receptor antagonists will be effective antipsychotic agents. In this regard, an amelioration of the negative and cognitive symptoms of schizophrenia holds the most promise for D(3) receptor antagonists, a concept currently under clinical evaluation. In addition, D(3) receptors could be involved in behavioral sensitization and the potential application of D(3) receptor antagonists in the treatment of drug abuse is undergoing intensive experimental investigation.

Degeneration of dopaminergic neurons in the substantia nigra of zitter mutant rat and protection by chronic intake of Vitamin E.

Dopaminergic cell death in the ventral and dorsal tiers of substantia nigra pars copmacta (SNc) and their prevention by anti-oxidant diet was immunohistochemically studied in the zitter mutant rats, which are characterized by abnormal metabolism of superoxide. Similar to previous reports, the number of SNc neurons in Nissl-stained section decreased with age. Tyrosine hydroxylase (TH) immunohistochemistry demonstrated that the dopaminergic neurons in the ventral tier of SNc degenerated early, whereas the dorsal tier gradually degenerated with age. Thus, the ventral tier dopaminergic neurons are affected first, but the dorsal tier neurons do become impact by the zi/zi mutation. Following 9-month period after weaning, zitter rats supplemented with 500 mg D,L-alpha-tocophenol (VE(+))/kg diet exhibited a significant increased of surviving TH-immunoreactive neurons in both the tiers of SNc as compared with the zi/zi rats with control and VE(-) diets. These results suggest that VE supplement may slow the dopaminergic cell loss in zitter mutant rat, and further support that degeneration of the dopaminergic neurons in this mutant rat is caused by oxidant stress. Thus, the zitter rat may represent a good model for studying the dopaminergic cell death by superoxide species.

Low dose pramipexole is neuroprotective in the MPTP mouse model of Parkinson's disease, and downregulates the dopamine transporter via the D3 receptor.

BACKGROUND: Our aim was to determine if pramipexole, a D3 preferring agonist, effectively reduced dopamine neuron and fiber loss in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model when given at intraperitoneal doses corresponding to clinical doses. We also determined whether subchronic treatment with pramipexole regulates dopamine transporter function, thereby reducing intracellular transport of the active metabolite of MPTP, 1-methyl-4-phenylpyridinium (MPP+). METHODS: Ten 12-month old C57BL/6 mice were treated with MPTP (or saline) twice per day at 20 mg/kg s.c. (4 injections over 48 h). Mice were pretreated for 3 days and during the 2-day MPTP regimen with pramipexole (0.1 mg/kg/day) or saline. Stereological quantification of dopamine neuron number and optical density measurement of dopamine fiber loss were carried out at 1 week after treatment, using immunostaining for dopamine transporter (DAT) and tyrosine hydroxylase (TH). Additional wild-type (WT) and D3 receptor knockout (KO) mice were treated for 5 days with pramipexole (0.1 mg/kg/day) or vehicle. The kinetics of [3H]MPP+ and [3H]DA uptake (Vmax and Km) were determined 24 h later; and at 24 h and 14 days dopamine transporter density was measured by quantitative autoradiography. RESULTS: Pramipexole treatment completely antagonized the neurotoxic effects of MPTP, as measured by substantia nigra and ventral tegmental area TH-immunoreactive cell counts. MPTP- induced loss of striatal innervation, as measured by DAT-immunoreactivity, was partially prevented by pramipexole, but not with regard to TH-IR. Pramipexole also reduced DAT- immunoreactivity in non-MPTP treated mice. Subchronic treatment with pramipexole lowered the Vmax for [3H]DA and [3H]MPP+ uptake into striatal synaptosomes of WT mice. Pramipexole treatment lowered Vmax in WT but not D3 KO mice; however, D3 KO mice had lower Vmax for [3H]DA uptake. There was no change in DAT number in WT with pramipexole treatment or D3 KO mice at 24 h post-treatment, but there was a reduction in WT-pramipexole treated and not in D3 KO mice at 14 days post-treatment. CONCLUSION: These results suggest that protection occurs at clinically suitable doses of pramipexole. Protection could be due to a reduced amount of MPP+ taken up into DA terminals via DAT. D3 receptor plays an important role in this regulation of transporter uptake and availability.

Substantia nigra Marinesco bodies are associated with decreased striatal expression of dopaminergic markers.

Marinesco bodies are nuclear inclusions found in pigmented neurons of the substantia nigra and locus ceruleus of humans and monkeys. It has long been known that the frequency of these inclusions increases with advancing age, but no pathologic associations have ever been established. We quantified Marinesco body frequency in human autopsy subjects, classified as young normal controls, elderly controls, dementia with Lewy bodies (DLB), Alzheimer disease (AD), and Parkinson disease (PD). Elderly controls, AD cases, and DLB cases had significantly increased Marinesco body frequencies relative to young controls and DLB cases had significantly increased frequencies relative to elderly controls, while PD cases did not differ from young controls; cases with AD did not differ from elderly controls. Lewy body-containing neurons had significantly higher Marinesco body frequencies than non-Lewy body-containing neurons. Marinesco body frequency in elderly control cases correlated significantly, in inverse fashion, with striatal concentrations of the dopaminergic neuron markers dopamine transporter and tyrosine hydroxylase. These statistical associations suggest that Marinesco bodies constitute or mark a pathologic process that may be related to, or contribute to, age-related motor dysfunction and/or Lewy body disorders. Further studies are needed to ascertain the molecular basis of Marinesco body formation; preliminary studies indicate that proteasome dysfunction can lead to similar abnormalities in cultured cells.

Increases in dopamine D3 receptor binding in rats receiving a cocaine challenge at various time points after cocaine self-administration: implications for cocaine-seeking behavior.

Previous research suggests that cocaine dysregulates dopamine D3 receptors. The present study examined the time course of changes in dopamine D3 receptor binding after terminating a cocaine self-administration regimen. [125I]-7-hydroxy-2-[N-propyl-N-(3'-iodo-2'-propenyl)-amino]-tetralin was used to label dopamine D3 receptors in rats that had undergone testing for cocaine-seeking behavior reinstated by a cocaine priming injection (15 mg/kg, i.p.; the behavior results have been previously published), and were killed 24 h after the test at time points that were either 2, 8, or 31-32 days after their last cocaine self-administration session. The results indicated a time-dependent increase in D3 receptor binding relative to controls that received saline yoked to the delivery of cocaine in an experimental animal. Specifically, there was no significant change in D3 receptor binding in cocaine-experienced rats killed at the 2- or 8-day time points relative to controls, but there was an increase in D3 receptor binding in the nucleus accumbens core and ventral caudate-putamen in rats killed at the 31- to 32-day time point. In a subsequent experiment, we replicated the increase in D3 receptor binding in rats that underwent a less extensive self-administration regimen, then were tested for cocaine-primed reinstatement of cocaine-seeking behavior, and then were killed 24 h later at a time point of 22 days after their last self-administration session. Furthermore, the increase in binding was attenuated by repeated 7-hydroxy-N,N-di-n-propyl-2-aminotetralin administration (1 mg/kg/day, s.c. for 14 days), a regimen that also reduces cocaine-seeking behavior in animals when tested in a nondrug state. Collectively, the findings suggest that regulatory responses of D3 receptors may be functionally related to changes in propensity for cocaine-seeking behavior.

Terminally differentiated SH-SY5Y cells provide a model system for studying neuroprotective effects of dopamine agonists.

We characterized undifferentiated (UN) and three differentiation conditions of the SH-SY5Y neuroblastoma cell line for phenotypic markers of dopaminergic cells, sensitivity to the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridinium ion (MPP+), the requirement to utilize the dopamine (DA) transporter (DAT) for MPP+ toxicity, and the neuroprotective effects of pramipexole. Cells were differentiated with retinoic acid (RA), 12-O-tetradecanoyl-phorbol-13-acetate (TPA), and RA followed by TPA (RA/TPA). RA/TPA treated cells exhibited the highest levels of tyrosine hydroxylase and DAT but lower levels of vesicular monoamine transporter. The kinetics of [3H]DA uptake and [3H]MPP+ uptake to DAT in RA/TPA differentiated cells were similar to that of rat and mouse caudate-putamen synaptosomes. RA/TPA differentiated cells evidenced high sensitivity to the neurotoxic effects of MPP+ (0.03 to 3.0 mM), and the neurotoxic effects of MPP+ were blocked with the DAT inhibitor 1-(2-[bis(4-fluorophenyl)methoxy]ethyl)-4-(3-phenylpropyl)piperazine (GBR 12909). DA-induced cell death was not more sensitive in RA vs RA/TPA differentiated cells and was not inhibited by transporter inhibitors. RA/TPA differentiated cells exhibited 3-fold and 6-fold higher levels, respectively, of DA D2 and D3 receptors than UN or RA differentiated cells. Pretreatment with pramipexole was protective against MPP+ in the RA/TPA differentiated cells but not in undifferentiated or RA differentiated cells. The neuroprotective effect of pramipexole was concentration-dependent and dopamine D2/D3 receptor dependent. In contrast, protection by pramipexole against DA was not DA receptor dependent. Further characterization of the neuroprotective effects of DA agonists in this model system can provide unique information about DA receptor dependent and independent mechanisms of neuroprotection.

Motor impairment in normal aging, clinically possible Parkinson's disease, and clinically probable Parkinson's disease: longitudinal evaluation of a cohort of prospective brain donors.

This study presents data on the antemortem evaluations of a cohort of individuals registered in a brain donation program. Clinical evaluation determined that many individuals were unaware they had clinical signs of Parkinson's disease (PD) (rest tremor, bradykinesia, rigidity). Quantitative motor testing (timed tapping test and Purdue pegboard test) revealed a graded reduction in performance in those clinically found to have clinically possible and clinically probable PD. Longitudinal examinations over 4 years revealed some individuals progressed from control to clinically possible PD and clinically possible PD to clinically probable PD. This study underscores the importance of longitudinal antemortem testing of prospective brain donors as well as the potential value of quantitative motor testing.

Behavioral effects of dopamine agonists and antagonists in MPTP-lesioned D3 receptor knockout mice.

To test the modulatory role of D(3) receptors in normal and dopamine-depleted mice, D(3) receptor KO mice and wild-type (WT) littermates were administered saline, L-dopa/carbidopa (20/2 mg/kg ip), a preferential D(3)>D(2) agonist S32504, a D1+D(2)/D(3) agonist apomorphine, a selective D(3) antagonist S33084, or apomorphine with S33084 prior to and after administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We monitored lines crossed in a 55-min session, average number of rears, and average number of grooming bouts. MPTP treatment produced equivalent 70% losses of dopamine fibers in the caudate putamen (CPu) and nucleus accumbens (NAC) of WT and D(3) KO mice as compared to their control (vehicle injected) counterparts. D(3) receptors were absent in KO mice, and the number of D(3) receptors was unaffected by MPTP-induced loss of DA terminals in WT mice. The results support a lack of involvement of the D(3) receptor for D1:D2 receptor-mediated behavioral activity (synergy). First, S32504 inhibited all behaviors and to a similar degree in D(3) KO and WT mice. Second, S33084 at the higher concentration increased number of lines crossed in response to high dose apomorphine in both D(3) KO and WT mice. Third, in nonlesioned mice, apomorphine-induced gnawing stereotypies were inhibited by S33084 in both D(3) KO and WT mice. Interestingly, the inhibition of apomorphine-induced gnawing was not apparent in MPTP-lesioned mice, and this stereotypy was elevated in D(3) KO-MPTP-lesioned mice. Thus, the suppressive effects of S32504 could be via D2 autoreceptor inhibition of DA release, and D2 receptor blockade by S33084 leads to release of that inhibition. This may be more apparent in MPTP-lesioned partially DA denervated mice.

Locomotor activity induced by MK-801 is enhanced in dopamine D3 receptor knockout mice but suppression by dopamine D3/D2 antagonists does not occur through the dopamine D3 receptor.

There are contradictory data regarding the role of the dopamine D(3) receptor in regulating N-methyl-d-aspartate (NMDA) receptor antagonist (e.g., dizocilpine) induced hyperactivity. The purpose of the present study was to examine the interaction of dopamine D(3) receptor preferring antagonists U99194A (5,6-dimethoxy-2(dipropylamino)indan) and S33804 ((3aR,9bS)-N[4-(8-cyano-1,3a,4,9b-tetrahydro-3H-benzopyrano[3,4-c]pyrrole-2-yl)-butyl] (4-phenyl)benzamide)) with dizocilpine (MK-801)-induced hyperactivity in wild type (WT) and dopamine D(3) receptor mutant (D(3)R KO) mice. D(3)R KO and WT mice were administered vehicle (saline, 1 ml/100g body weight, i.p.), or S33084 (1.0mg/kg.) and U99194A (0.1mg/kg or 0.01 mg/kg), and horizontal and vertical activity counts were recorded for 30 min. Mice were then treated with vehicle or MK-801 (0.12 mg/kg i.p.) and returned to the open field for an additional 55 min. D(3)R KO mice showed a significantly higher level of locomotor and rearing activity during the 1st 30 min after vehicle treatment compared to WT mice. MK-801-hyperactivity was significantly higher in D(3)R KO mice than WT mice. Dopamine D(3) receptor preferring antagonists suppressed the locomotor activity response to MK-801 to an equal extent in D(3)R KO and WT mice. The results confirm that MK-801-induced hyperactivity and novelty-induced behavioral activity and rearing are enhanced in D(3)R KO mice, but suppression by dopamine D(3) receptor preferring antagonists is not through dopamine D(3) receptor antagonism.

Age-related differences in MK-801 induced behaviors in dopamine D3 receptor knock out mice.

It is not known if age plays an important role in the D(3) receptor regulation of N-methyl-D-aspartate (NMDA) receptor antagonist induced hyperactivity. Wild type (WT) and dopamine D(3) receptor mutant (D(3)R KO) mice were divided into young (under 7 months) and middle age (over 12 months) groups and tested for dizocilpine (MK-801)-induced hyperactivity and rearing. Mice were administered vehicle (saline, 1 ml/100g body weight, i.p.), or dopamine D(3) receptor preferring antagonists 3aR,9bS)-N[4-(8-cyano-1,3a,4,9b-tetrahydro-3H-benzopyrano[3,4-c]pyrrole-2-yl)-butyl] (4-phenyl) benzamide) (S33084, 1.0mg/kg, i.p.) and 5,6-dimethoxy-2(dipropylamino)indan (U99194A, 5.0 mg/kg i.p.), and immediately placed into the open field apparatus. Horizontal and vertical activity counts were recorded for 30 min, followed by injection of vehicle or MK801 (0.15 or 0.30 mg/kg i.p.) and mice returned to the open field for an additional 55 min. Young D(3)R KO mice showed the highest level of locomotor and rearing activity during the 1st 30 min and 2nd 55 min session after vehicle treatment. At the lower dose of MK-801 horizontal activity was significantly higher in Young-D(3)R KO mice than in the other groups. At the higher dose of MK-801 horizontal activity was elevated to an equal extent in all groups. In response to S33084 and U99194A, MK-801 hyperactivity was reduced the most in the Middle Age-D(3)R KO and the least in the Young-D(3)R KO mice. Rearing showed pronounced age-related but not genotype effects. The results demonstrate that MK-801 induced-hyperactivity, novelty-induced behavioral activity and rearing are affected by age and D(3) receptor genotype.

Elevated tauopathy and alpha-synuclein pathology in postmortem Parkinson's disease brains with and without dementia.

Parkinson's disease (PD), a progressive neurodegenerative disease, results in abnormal accumulation of insoluble alpha-synuclein (alpha-Syn) in dopaminergic neurons. Here we examined tauopathic changes and the alpha-Syn/p-GSK-3beta/proteasome pathway in postmortem striata and inferior frontal gyri (IFG) from patients with PD and PD with dementia (PDD). In both PD and PDD, alpha-Syn levels were high, especially the insoluble form of this protein; in PDD, insoluble alpha-Syn levels were persistently higher than PD across both brain regions. Levels of p-GSK-3beta phosphorylated at Tyr 216, which hyperphosphorylates Tau to produce toxic pathological forms of p-Tau, were higher in striata of both PD and PDD compared to controls, but were unaltered in IFG. While proteasomal activity was unchanged in striatum of PD and PDD, such activity was diminished in the IFG of both PD and PDD. A decrease in 19S subunit of the proteasomes was seen in IFG of PDD, while lower levels of 20S subunits were seen in striatum and IFG of both PD and PDD patients. Parkin levels were similar in PD and PDD, suggesting lack of involvement of this protein. Most interestingly, tauopathic changes were noted only in striatum of PD and PDD, with increased hyperphosphorylation seen at Ser262 and Ser396/404; increases in Ser202 levels were seen only in PD but not in PDD striatum. We were unable to detect any tauopathy in IFG in either PD or PDD despite increased levels of alpha-Syn, and decreased proteasomal activity, and is probably due to lack of increase in p-GSK-3beta in IFG. Unlike Alzheimer's disease where tauopathy is more globally observed in diverse brain regions, our data demonstrates restricted expression of tauopathy in brains of PD and PDD, probably limited to dopaminergic neurons of the nigrostriatal region.

Small molecule BDNF mimetics activate TrkB signaling and prevent neuronal degeneration in rodents.

Brain-derived neurotrophic factor (BDNF) activates the receptor tropomyosin-related kinase B (TrkB) with high potency and specificity, promoting neuronal survival, differentiation, and synaptic function. Correlations between altered BDNF expression and/or function and mechanism(s) underlying numerous neurodegenerative conditions, including Alzheimer disease and traumatic brain injury, suggest that TrkB agonists might have therapeutic potential. Using in silico screening with a BDNF loop-domain pharmacophore, followed by low-throughput in vitro screening in mouse fetal hippocampal neurons, we have efficiently identified small molecules with nanomolar neurotrophic activity specific to TrkB versus other Trk family members. Neurotrophic activity was dependent on TrkB and its downstream targets, although compound-induced signaling activation kinetics differed from those triggered by BDNF. A selected prototype compound demonstrated binding specificity to the extracellular domain of TrkB. In in vitro models of neurodegenerative disease, it prevented neuronal degeneration with efficacy equal to that of BDNF, and when administered in vivo, it caused hippocampal and striatal TrkB activation in mice and improved motor learning after traumatic brain injury in rats. These studies demonstrate the utility of loop modeling in drug discovery and reveal what we believe to be the first reported small molecules derived from a targeted BDNF domain that specifically activate TrkB.We propose that these compounds constitute a novel group of tools for the study of TrkB signaling and may provide leads for developing new therapeutic agents for neurodegenerative diseases.

Microglial responses to dopamine in a cell culture model of Parkinson's disease.

Activated microglia appear to selectively attack dopamine (DA) neurons in the Parkinson's disease (PD) substantia nigra. We investigated potential mechanisms using culture models. As targets, human SH-SY5Y cells were left undifferentiated (UNDIFF) or were differentiated with retinoic acid (RA) or RA plus brain-derived neurotrophic factor (RA/BDNF). RA/BDNF-treated cells were immunoreactive for tyrosine hydroxylase and the DA transporter, took up exogenous DA, and released DA after K(+) stimulation. Undifferentiated and RA-treated cells lacked these characteristics of a DA phenotype. Co-culture of target cells with human elderly microglia resulted in elevated toxicity in DA phenotype (RA/BDNF) cells. Lipopolysaccharide (LPS) plus K(+)-stimulated DA release enhanced toxicity by 500-fold. DA induced microglial chemotaxis in Boyden chambers. Spiperone inhibited this effect. Cultured human elderly microglia expressed mRNAs for D1-D4 but not D5 DA receptors. The microglia, as well as PD microglia in situ, were also immunoreactive for D1-D4 but not D5 DA receptors. These findings demonstrate that activated microglia express DA receptors, and suggest that this mechanism may play a role in the selective vulnerability of DA neurons in PD.

Upregulation of Arc mRNA expression in the prefrontal cortex following cue-induced reinstatement of extinguished cocaine-seeking behavior.

Cocaine-associated cues acquire incentive motivational effects that manifest as cue-elicited craving in humans and cocaine-seeking behavior in rats. Here we examine the hypothesis that neuronal processes associated with incentive motivational effects of cocaine cues involve increased expression of the plasticity-associated gene, Arc. Rats trained to self-administer cocaine subsequently underwent extinction training, during which cocaine-seeking behavior (i.e., responses without cocaine reinforcement) progressively decreased. Rats were then tested for cocaine-seeking behavior either with or without response-contingent presentations of light/tone cues that had been previously paired with cocaine infusions during self-administration training. Cues elicited reinstatement of cocaine-seeking behavior and were accompanied by increased Arc mRNA levels in the orbitofrontal, prelimbic, and anterior cingulate cortices, suggesting Arc involvement in conditioned plasticity associated with incentive motivational effects of cocaine cues. Additionally, rats with a history of cocaine self-administration and extinction exhibited upregulation of Arc expression in several limbic and cortical regions relative to saline-yoked controls regardless of cue exposure condition, suggesting persistent neuroadaptations involving Arc within these regions.