Anna-Liisa Brownell

Embryonic stem cells develop into functional dopaminergic neurons after transplantation in a Parkinson rat model

Although implantation of fetal dopamine (DA) neurons can reduce parkinsonism in patients, current methods are rudimentary, and a reliable donor cell source is lacking. We show that transplanting low doses of undifferentiated mouse embryonic stem (ES) cells into the rat striatum results in a proliferation of ES cells into fully differentiated DA neurons. ES cell-derived DA neurons caused gradual and sustained behavioral restoration of DA-mediated motor asymmetry. Behavioral recovery paralleled in vivo positron emission tomography and functional magnetic resonance imaging data demonstrating DA-mediated hemodynamic changes in the striatum and associated brain circuitry. These results demonstrate that transplanted ES cells can develop spontaneously into DA neurons. Such DA neurons can restore cerebral function and behavior in an animal model of Parkinsons disease.

Systemic exposure to proteasome inhibitors causes a progressive model of Parkinson's disease

Environmental toxins have been implicated in the etiology of Parkinsons disease. Recent findings of defects in the ubiquitin‐proteasome system in hereditary and sporadic forms of the illness suggest that environmental proteasome inhibitors are candidate PD‐inducing toxins. Here, we systemically injected six doses of naturally occurring (epoxomicin) or synthetic (Z‐lle‐Glu(OtBu)‐Ala‐Leu‐al [PSI]) proteasome inhibitors into adult rats over a period of 2 weeks. After a latency of 1 to 2 weeks, animals developed progressive parkinsonism with bradykinesia, rigidity, tremor, and an abnormal posture, which improved with apomorphine treatment. Positron emission tomography demonstrated reduced carbon‐11‐labeled 2β‐carbomethoxy‐3β‐(4‐fluorophenyl)tropane (CFT) binding to dopaminergic nerve terminals in the striatum, indicative of degeneration of the nigrostriatal pathway. Postmortem analyses showed striatal dopamine depletion and dopaminergic cell death with apoptosis and inflammation in the substantia nigra pars compacta. In addition, neurodegeneration occurred in the locus coeruleus, dorsal motor nucleus of the vagus, and the nucleus basalis of Meynert. At neurodegenerative sites, intracytoplasmic, eosinophilic, α‐synuclein/ubiquitin–containing, inclusions resembling Lewy bodies were present in some of the remaining neurons. This animal model induced by proteasome inhibitors closely recapitulates key features of PD and may be valuable in studying etiopathogenic mechanisms and putative neuroprotective therapies for the illness. Ann Neurol 2004

Neuroinflammation of the nigrostriatal pathway during progressive 6-OHDA dopamine degeneration in rats monitored by immunohistochemistry and PET imaging

We investigated the microglial response to progressive dopamine neuron degeneration using in vivo positron emission tomography (PET) imaging and postmortem analyses in a Parkinsons disease (PD) rat model induced by unilateral (right side) intrastriatal administration of 6‐hydroxydopamine (6‐OHDA). Degeneration of the dopamine system was monitored by PET imaging of presynaptic dopamine transporters using a specific ligand 11C‐CFT (2β‐carbomethoxy‐3β‐(4‐fluorophenyl) tropane). Binding of 11C‐CFT was markedly reduced in the striatum indicating dopaminergic degeneration. Parallel PET studies of 11C‐PK11195 (1‐(2‐chlorophenyl)‐N‐methyl‐N‐(1‐methylpropyl)‐3 isoquinoline carboxamide) (specific ligand for activated microglia) showed increased binding in the striatum and substantia nigra indicative of a microglial response. Postmortem immunohistochemical analyses were performed with antibodies against CR3 for microglia/macrophage activation. Using a qualitative postmortem index for microglial activation we found an initially focal, then widespread microglial response at striatal and nigral levels at 4 weeks postlesion. These data support the hypothesis that inflammation is a significant component of progressive dopaminergic degeneration that can be monitored by PET imaging.

Selective COX-2 inhibition prevents progressive dopamine neuron degeneration in a rat model of Parkinson's disease

Several lines of evidence point to a significant role of neuroinflammation in Parkinsons disease (PD) and other neurodegenerative disorders. In the present study we examined the protective effect of celecoxib, a selective inhibitor of the inducible form of cyclooxygenase (COX-2), on dopamine (DA) cell loss in a rat model of PD. We used the intrastriatal administration of 6-hydroxydopamine (6-OHDA) that induces a retrograde neuronal damage and death, which progresses over weeks. Animals were randomized to receive celecoxib (20 mg/kg/day) or vehicle starting 1 hour before the intrastriatal administration of 6-OHDA. Evaluation was performed in vivo using micro PET and selective radiotracers for DA terminals and microglia. Post mortem analysis included stereological quantification of tyrosine hydroxylase, astrocytes and microglia. 12 days after the 6-OHDA lesion there were no differences in DA cell or fiber loss between groups, although the microglial cell density and activation was markedly reduced in animals receiving celecoxib (p < 0.01). COX-2 inhibition did not reduce the typical astroglial response in the striatum at any stage. Between 12 and 21 days, there was a significant progression of DA cell loss in the vehicle group (from 40 to 65%) that was prevented by celecoxib. Therefore, inhibition of COX-2 by celecoxib appears to be able, either directly or through inhibition of microglia activation to prevent or slow down DA cell degeneration.

Successful Function of Autologous iPSC-Derived Dopamine Neurons following Transplantation in a Non-Human Primate Model of Parkinson’s Disease

Autologous transplantation of patient-specific induced pluripotent stem cell (iPSC)-derived neurons is a potential clinical approach for treatment of neurological disease. Preclinical demonstration of long-term efficacy, feasibility, and safety of iPSC-derived dopamine neurons in non-human primate models will be an important step in clinical development of cell therapy. Here, we analyzed cynomolgus monkey (CM) iPSC-derived midbrain dopamine neurons for up to 2 years following autologous transplantation in a Parkinsons disease (PD) model. In one animal, with the most successful protocol, we found that unilateral engraftment of CM-iPSCs could provide a gradual onset of functional motor improvement contralateral to the side of dopamine neuron transplantation, and increased motor activity, without a need for immunosuppression. Postmortem analyses demonstrated robust survival of midbrain-like dopaminergic neurons and extensive outgrowth into the transplanted putamen. Our proof of concept findings support further development of autologous iPSC-derived cell transplantation for treatment of PD.

DIAGNOSIS OF ACUTE HERPES SIMPLEX ENCEPHALITIS BY BRAIN PERFUSION SINGLE PHOTON EMISSION COMPUTED TOMOGRAPHY

Brain perfusion was studied in 14 patients with acute encephalitis by use of 123I-iodoamphetamine or 99mTc-hexamethylpropyleneamine oxime and single photon emission computed tomography (SPECT), the first examination being made 4-11 days after onset of encephalitis symptoms. All 6 patients with herpes simplex virus encephalitis (HSVE) had strongly increased accumulation of radiotracer in the affected temporal lobe; in the remaining 8 results were normal. At the time of the first SPECT conventional CT images were normal in all patients. The SPECT abnormality in HSVE gradually converted over 4-10 weeks from increased tracer accumulation to greatly subnormal accumulation. Brain perfusion SPECT may be helpful in the early diagnosis of HSVE.

Detection of the effects of dopamine receptor supersensitivity using pharmacological MRI and correlations with PET

Receptor supersensitivity is an important concept for understanding neurotransmitter and receptor dynamics. Traditionally, detection of receptor supersensitivity has been performed using autoradiography or positron emission tomography (PET). We show that use of magnetic resonance imaging (MRI) not only enables one to detect dopaminergic supersensitivity, but that the hemodynamic time course reflective of this fact is different in different brain regions. In rats unilaterally lesioned with intranigral 6‐hydroxydopamine, apomorphine injections lead to a large increase in hemodynamic response (cerebral blood volume, CBV) in the striato‐thalamo‐cortico circuit on the lesioned side but had little effect on the intact side. Amphetamine injections lead to increases in hemodynamic responses on the intact side and little on the lesioned side in the same animals. The time course for the increase in CBV after either amphetamine or apomorphine administration was longer in striatum and thalamus than in frontal cortex. 11C‐PET studies of ligands which bind to the dopamine transporter (2‐β‐carbomethoxy‐3‐β‐(4‐fluorophenyl)tropane 1,5‐naphthalnendisulfonate, WIN 35, 428 or CFT) and D2 receptors (raclopride) confirm that there is a loss of presynaptic dopamine terminals as well as upregulation of D2 receptors in striatum in these same animals. Pharmacologic MRI should become a sensitive tool to measure functional supersensitivity in humans, providing a complementary picture to that generated using PET studies of direct receptor binding. Synapse 36:57–65, 2000.

Dopamine fiber detection by [11c]-cft and Pet in a primate model of parkinsonism

MONKEYS were treated on two regimens of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) injections to achieve dopamine fiber degeneration of differing severities. A rapid treatment regimen produced a severe parkinsonian syndrome, whereas an intermittent regimen did not cause locomotor symptoms to appear up to 25 weeks. High resolution PET scanning of dopamine nerve terminals revealed that the specific binding of the dopamine transporter [11C]-WIN 35,428 ([11C]-CFT) was diminished by 94% (caudate nucleus) and by 93% (putamen) in the symptomatic monkey. Decreases of 65 and 67% were detected in these regions in the non-symptomatic monkey. Post-mortem immunocytochemical evaluation of presumed dopamine fibers by tyrosine-hydroxylase showed similar reductions in the symptomatic animal.

Combined PET/MRS brain studies show dynamic and long-term physiological changes in a primate model of Parkinson disease

We used brain imaging to study long-term neurodegenerative and bioadaptive neurochemical changes in a primate model of Parkinson disease. We gradually induced a selective loss of nigrostriatal dopamine neurons, similar to that of Parkinson disease, by creating oxidative stress through infusion of the mitochondrial complex 1 inhibitor MPTP for 14 ± 5 months. Repeated evaluations over 3 years by positron emission tomography (PET) demonstrated progressive and persistent loss of neuronal dopamine pre-synaptic re-uptake sites; repeated magnetic resonance spectroscopy (MRS) studies indicated a 23-fold increase in lactate and macromolecules in the striatum region of the brain for up to 10 months after the last administration of MPTP. By 2 years after the MPTP infusions, these MRS striatal lactate and macromolecule values had returned to normal levels. In contrast, there were persistent increases in striatal choline and decreases in N-acetylaspartate. Thus, these combined PET/MRS studies demonstrate patterns of neurochemical changes that are both dynamic and persistent long after selective dopaminergic degeneration.

Mapping Dopamine Function in Primates Using Pharmacologic Magnetic Resonance Imaging

Dopamine (DA) receptors play a central role in such diverse pathologies as Parkinsons disease, schizophrenia, and drug abuse. We used an amphetamine challenge combined with pharmacologic magnetic resonance imaging (phMRI) to map DA-associated circuitry in nonhuman primates with high sensitivity and spatial resolution. Seven control cynomolgous monkeys and 10 MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine)-treated parkinsonian primates were studied longitudinally using both positron emission tomography (PET) and phMRI. Amphetamine challenge (2.5 mg/kg, i.v.) in control monkeys increased relative cerebral blood volume (rCBV) in a number of brain regions not described previously, such as parafascicular thalamus, precentral gyrus, and dentate nucleus of the cerebellum. With the high spatial resolution, we were also able to readily identify changes in rCBV in the anterior cingulate, substantia nigra, ventral tegmental area, caudate (tail and head), putamen, and nucleus accumbens. Amphetamine induced decreases in rCBV in occipital and posterior parietal cortices. Parkinsonian primates had a prominent loss of response to amphetamine, with relative sparing of the nucleus accumbens and parafascicular thalamus. There was a significant correlation between rCBV loss in the substantia nigra and both PET imaging of dopamine transporters and behavioral measures. Monkeys with partial lesions as defined by 2β-carbomethoxy-3β-(4-fluorophenyl) tropane binding to dopamine transporters showed recruitment of premotor and motor cortex after amphetamine stimulus similar to what has been noted in Parkinsons patients during motor tasks. These data indicate that phMRI is a powerful tool for assessment of dynamic changes associated with normal and dysfunctional DA brain circuitry in primates.