Sylvia S. White

Compensatory Changes in the Noradrenergic Nervous System in the Locus Ceruleus and Hippocampus of Postmortem Subjects with Alzheimer's Disease and Dementia with Lewy Bodies

In Alzheimers disease (AD), there is a significant loss of locus ceruleus (LC) noradrenergic neurons. However, functional and anatomical evidence indicates that the remaining noradrenergic neurons may be compensating for the loss. Because the noradrenergic system plays an important role in learning and memory, it is important to determine whether compensation occurs in noradrenergic neurons in the LC and hippocampus of subjects with AD or a related dementing disorder, dementia with Lewy bodies (DLB). We observed profound neuronal loss in the LC in AD and DLB subjects with three major changes in the noradrenergic system consistent with compensation: (1) an increase in tyrosine hydroxylase (TH) mRNA expression in the remaining neurons; (2) sprouting of dendrites into peri-LC dendritic zone, as determined by α2-adrenoreceptors (ARs) and norepinephrine transporter binding sites; and (3) sprouting of axonal projections to the hippocampus as determined by α2-ARs. In AD and DLB subjects, the postsynaptic α1-ARs were normal to elevated. Expression of α1A- and α2A-AR mRNA in the hippocampus of AD and DLB subjects were not altered, but expression of α1D- and α2C-AR mRNA was significantly reduced in the hippocampus of AD and DLB subjects. Therefore, in AD and DLB subjects, there is compensation occurring in the remaining noradrenergic neurons, but there does appear to be a loss of specific AR in the hippocampus. Because changes in these noradrenergic markers in AD versus DLB subjects were similar (except neuronal loss and the increase in TH mRNA were somewhat greater in DLB subjects), the presence of Lewy bodies in addition to plaques and tangles in DLB subjects does not appear to further affect the noradrenergic compensatory changes.

Differential response of the central noradrenergic nervous system to the loss of locus coeruleus neurons in Parkinson's disease and Alzheimer's disease.

In Parkinsons disease (PD), there is a significant loss of noradrenergic neurons in the locus coeruleus (LC) in addition to the loss of dopaminergic neurons in the substantia nigra (SN). The goal of this study was to determine if the surviving LC noradrenergic neurons in PD demonstrate compensatory changes in response to the neuronal loss, as observed in Alzheimers disease (AD). Tyrosine hydroxylase (TH) and dopamine β-hydroxylase (DBH) mRNA expression in postmortem LC tissue of control and age-matched PD subjects demonstrated a significant reduction in the number of noradrenergic neurons in the LC of PD subjects. TH mRNA expression/neuron did not differ between control and PD subjects, but DBH mRNA expression/neuron was significantly elevated in PD subjects compared to control. This increase in DBH mRNA expression in PD subjects is not a response to neuronal loss because the amount of DBH mRNA expression/neuron in AD subjects was not significantly different from control. Norepinephrine transporter (NET) binding site concentration in the LC of PD subjects was significantly reduced over the cell body region as well as the peri-LC dendritic zone. In PD subjects, the loss of dendrites from surviving noradrenergic neurons was also apparent with TH-immunoreactivity (IR). This loss of LC dendritic innervation in PD subjects as measured by TH-IR was not due to LC neuronal loss because TH-IR in AD subjects was robust, despite a similar loss of LC neurons. These data suggest that there is a differential response of the noradrenergic nervous system in PD compared to AD in response to the loss of LC neurons.

A Comprehensive Analysis of the Effect of DSP4 on the Locus Coeruleus Noradrenergic System in the Rat

Degeneration of the noradrenergic neurons in the locus coeruleus (LC) is a major component of Alzheimers (AD) and Parkinsons disease (PD), but the consequence of noradrenergic neuronal loss has different effects on the surviving neurons in the two disorders. Therefore, understanding the consequence of noradrenergic neuronal loss is important in determining the role of this neurotransmitter in these neurodegenerative disorders. The goal of the study was to determine if the neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP4) could be used as a model for either (or both) AD or PD. Rats were administered DSP4 and sacrificed 3 days 2 weeks and 3 months later. DSP4-treatment resulted in a rapid, though transient reduction in norepinephrine (NE) and NE transporter (NET) in many brain regions receiving variable innervation from the LC. Alpha(1)-adrenoreceptors binding site concentrations were unchanged in all brain regions at all three time points. However, an increase in alpha(2)-AR was observed in many different brain regions 2 weeks and 3 months after DSP4. These changes observed in forebrain regions occurred without a loss in LC noradrenergic neurons. Expression of synthesizing enzymes or NET did not change in amount of expression/neuron despite the reduction in NE tissue content and NET binding site concentrations at early time points, suggesting no compensatory response. In addition, DSP4 did not affect basal activity of LC at any time point in anesthetized animals, but 2 weeks after DSP4 there is a significant increase in irregular firing of noradrenergic neurons. These data indicate that DSP4 is not a selective LC noradrenergic neurotoxin, but does affect noradrenergic neuron terminals locally, as evident by the changes in transmitter and markers at terminal regions. However, since DSP4 did not result in a loss of noradrenergic neurons, it is not considered an adequate model for noradrenergic neuronal loss observed in AD and PD.

Regulation of norepinephrine transporter abundance by catecholamines and desipramine in vivo

The norepinephrine transporter (NET) regulates adrenoreceptor signaling by controlling the availability of synaptic norepinephrine (NE), and it is a direct target for some classes of antidepressant drugs. NET levels are normal in dopamine beta-hydroxylase knockout (Dbh -/-) mice that lack NE, demonstrating that the NET does not require endogenous NE for appropriate regulation under physiological conditions. In contrast, tyrosine hydroxylase knockout (Th -/-) mice that lack both NE and dopamine (DA) have reduced levels of NET, suggesting that it is down-regulated by a complete absence of catecholamines and not NE per se. Chronic treatment with the NET inhibitor, desipramine (DMI), reduced NET levels in both control and Dbh -/- mice, demonstrating that NE is not required for the regulation of NET by antidepressant drugs. There are some qualitative and quantitative differences in the down-regulation of the NET by catecholamine depletion and DMI treatment, suggesting that different mechanisms may be involved.

Changes in adrenoreceptors in the prefrontal cortex of subjects with dementia: Evidence of compensatory changes

In Alzheimers disease (AD) there is a significant loss of locus coeruleus (LC) noradrenergic neurons. However, recent work has shown the surviving noradrenergic neurons to display many compensatory changes, including axonal sprouting to the hippocampus. The prefrontal cortex (PFC) is a forebrain region that is affected in dementia, and receives innervation from the LC noradrenergic neurons. Reduced PFC function can reduce cognition and disrupt behavior. Because the PFC is an important area in AD, we determined if noradrenergic innervation from the LC noradrenergic neurons is maintained and if adrenoreceptors are altered postsynaptically. Presynaptic PFC alpha2-adrenoreceptor (AR) binding site density, as determined by 3H-RX821002, suggests that axons from surviving noradrenergic neurons in the LC are sprouting to the PFC of subjects with dementia. Changes in postsynaptic alpha1-AR in the PFC of subjects with dementia indicate normal to elevated levels of binding sites. Expression of alpha1-AR subtypes (alpha1A- and alpha1D-AR) and alpha2C-AR subtype mRNA in the PFC of subjects with dementia is similar to what was observed in the hippocampus with one exception, the expression of alpha1A-AR mRNA. The expression of the alpha1A-AR mRNA subtype is significantly reduced in specific layers of the PFC in subjects with dementia. The loss of alpha1A-, alpha1D- and alpha2C-AR mRNA subtype expression in the PFC may be attributed to neuronal loss observed in dementia. These changes in postsynaptic AR would suggest a reduced function of the PFC. Consequence of this reduced function of the PFC in dementia is still unknown but it may affect memory and behavior.

Tyrosine hydroxylase mRNA is increased in old age and norepinephrine uptake transporter mRNA is decreased in middle age in locus coeruleus of Brown-Norway rats.

In normal aging, cell loss occurs in the locus coeruleus (LC), the major noradrenergic nucleus in the brain. This study examined changes in the LC of aged rats by measuring mRNA expression for tyrosine hydroxylase (TH) and the norepinephrine uptake transporter (NET). TH and NET mRNA expression were measured by in situ hybridization in young, middle-aged and aged rats. It appears that in middle age, the transporter system responds initially to LC cell loss by decreasing NET mRNA expression. Then, with further aging and cell loss, TH mRNA expression increases which may potentially increase NE synthesis in the remaining neurons. These findings suggest that multiple regulatory components are used to maintain stable noradrenergic synaptic levels despite neuronal loss. Published by Elsevier Science B.V.

Effect of pentylenetetrazol on the expression of tyrosine hydroxylase mRNA and norepinephrine and dopamine transporter mRNA

Seizure activity has been shown to have differential effects on the terminal content of the monoamines, norepinephrine (NE) and dopamine (DA). Induction of seizure activity reduces the terminal content of NE, while DA levels remain unchanged or slightly elevated. This study examined the effect of the chemoconvulsant pentylenetetrazol (PTZ) on the mRNA expression of regulatory proteins which maintain the terminal content of NE and DA (i.e., synthesis and re-uptake). The areas examined were the noradrenergic neurons of the locus coeruleus (LC) and dopaminergic neurons of the substantia nigra pars compacta/ventral tegmentum area (SNpc/VTA) in the rat. In the LC, PTZ increased mRNA expression of the immediate early gene, c-fos, and mRNA expression of the synthesizing enzyme, tyrosine hydroxylase (TH), and the re-uptake protein, norepinephrine transporter (NET). This effect on TH and NET was observed only 1 day after the administration of PTZ. In contrast, PTZ did not alter the expression of c-fos mRNA in the SNpc/VTA, but reduced the expression of the dopamine transporter (DAT) mRNA. This effect was observed only 1 day after the administration of PTZ. TH mRNA expression in dopaminergic neurons was elevated initially in a manner similar to that observed in the LC. However, the effect of PTZ on TH mRNA expression in dopaminergic neurons was more prolonged (still elevated 3 days later). These results indicate that the chemoconvulsant PTZ has differential effects on the mRNA expression of regulatory systems (TH and neurotransporter proteins) in noradrenergic and dopaminergic neurons.

Alterations in mRNA expression of systems that regulate neurotransmitter synaptic content in seizure-naive genetically epilepsy-prone rat (GEPR): transporter proteins and rate-limiting synthesizing enzymes for norepinephrine, dopamine and serotonin.

Two models of genetically epilepsy-prone rat (GEPR) exist, the GEPR-3 and GEPR-9, GEPR-3 and GEPR-9 share a deficiency in presynaptic norepinephrine (NE) and serotonin (5HT) content in specific regions of the central nervous system (CNS). The presynaptic content of dopamine (DA) does not appear to be altered in either adult GEPR strain compared to Sprague-Dawley (SD) rats, the strain from which the GEPR was derived. Presynaptic content of monoamine neurotransmitters, such as NE, 5HT and DA, are maintained by several regulatory proteins which include: synthesis, re-uptake, release, degradation and vesicular transport. To further characterize the monoamine deficiency observed in the GEPR, the mRNA level of the rate limiting enzymes for the synthesis of NE, 5HT and DA and each of the neurotransporter proteins were measured in seizure-naive GEPR-3, GEPR-9 and SD rats. In the locus coeruleus (LC), the major noradrenergic locus, tyrosine hydroxylase (TH) mRNA level was significantly reduced only in GEPR-9 animals compared to SD rats and GEPR-3, while NE transporter (NET) mRNA was significantly elevated in GEPR-3 compared to SD rats and GEPR-9. TH and DA transporter (DAT) mRNA was measured in the dopaminergic neurons of the substantia nigra pars compacta (SNpc), ventral tegmental area (VTA) and zona incerta (ZI), DAT mRNA level was significantly reduced in all dopaminergic neurons in the GEPR-3 compared to SD rats and GEPR-9, while TH mRNA level was significantly elevated in the SNpc/VTA equally in GEPR-3 and GEPR-9 compared to SD rats. In the ZI, TH mRNA level was significantly reduced in GEPR-3 compared to SD rats and GEPR-9. In the dorsal raphe (DR), a major serotonergic locus, tryptophan hydroxylase (TRH) mRNA level was not significantly different from SD in either strain of GEPR; however, 5HT transporter (SERT) mRNA level was significantly reduced in GEPR-9 in the dorsal and lateral regions of the DR compared in SD rats and GEPR-3. These data indicate that two of the regulatory systems that maintain NE, 5HT and DA content are altered in a differential manner in seizure-naive GEPR-3 compared to seizure-naive GEPR-9, with GEPR-3 showing more alterations in dopaminergic neurons. It is uncertain at the present time how these alterations in mRNA level relate to the enhanced seizure susceptibility of these animals. It was apparent that a straightforward correlation between neurotransmitter loss to transcriptional changes in synthesizing enzymes mRNA or to re-uptake protein mRNA was not observed in noradrenergic and serotonergic neurons. Therefore, the decrease in presynaptic NE and 5HT tissue content in these animals may be due to posttranscriptional modification. In contrast, presynaptic DA tissue content which was unaltered in both strains of GEPR, shows an alteration in TH and DAT mRNA level compared to SD rats in all dopaminergic neurons examined. This indicates a possible involvement of DA in regulating the seizure susceptibility of these animals.

The ketogenic diet does not alter brain expression of orexigenic neuropeptides

Neuropeptide Y (NPY) and galanin are neuropeptides that are regulated by energy states and are also anticonvulsant. We tested the hypothesis that the anticonvulsant efficacy of the ketogenic diet (KD) is mediated by increased expression of NPY and galanin via alterations in food intake and energy metabolism. In situ hybridization revealed no effect of the KD on NPY or galanin mRNA expression, suggesting that increased expression of NPY and galanin do not contribute to the anticonvulsant effect of the KD.

Behavioral and metabolic features of repetitive seizures in immature and mature rats

Seizure incidence varies significantly with age, with seizure susceptibility particularly high during the first few years of life. Of significant concern is what effects do brief, repetitive seizures have on the developing brain. We approached this issue by examining the change in seizure threshold, and related markers of neuronal activity and metabolic activity (c-fos mRNA and 2-deoxyglucose [2DG]), as a function of repetitive seizure episodes in immature and mature rats. Starting on postnatal day 15 (P15) (immature) or P60 (adult) rats were given two flurothyl seizures a day for 5 days (nine or ten seizures). The seizure latency profile, our measure of threshold, in immature versus adult rats across the 5-day testing period was different. In immature rats, threshold for the second seizure on each day was significantly lower than for the first seizure, suggesting that there was little refractoriness after the first seizure of the day. In contrast, the mature animal had a significantly longer threshold latency to the second seizure for the first 3 days of testing. The immature animal was also more likely than the adult to exhibit tonic extension as a feature of the first seizure of the day. Following repetitive seizures, more regions of the CNS showed c-fos mRNA expression in the immature animal than adults, suggesting that repetitive seizures in the immature animal activated a greater percentage of the brain. Compared with the effects of a single seizure, repetitive seizures resulted in less 2DG labeling in most regions of the brain (except the hippocampus); in the immature brain this difference was more distinct than in adults. The consequences of repetitive seizures in the immature animal results in distinctly different seizure behavior and neuronal activity pattern (c-fos expression) than that observed in the mature animal.