The spleen: the forgotten organ in acute kidney injury of critical illness.

Acute kidney injury (AKI) is an increasing medical burden and is independently associated with mortality. AKI is a common comorbidity in the intensive care unit (ICU), with sepsis-associated AKI seen in almost a quarter of all ICU patients. Due to the high mortality seen in these patients, improved therapeutic options are needed. Data from experimental studies in animals support observations in humans that the host immune response to sepsis and trauma contributes to multiorgan failure and the high morbidity and mortality seen in critically ill patients. The spleen, a major component of the reticuloendothelial system, appears to be a key player in the 'cytokine storm' that develops after infection and trauma, and the resultant systemic inflammation is regulated by the autonomic nervous system. Over the past decade, evidence has suggested that controlling the splenic cytokine response improves tissue function and mortality in sepsis and other inflammatory-mediated diseases. One pathway that controls the response of the spleen to sepsis and trauma is the cholinergic anti-inflammatory pathway, and it may provide a key target for therapeutic intervention. Here, we review this concept and highlight the potential use of ultrasound to stimulate the cholinergic anti-inflammatory pathway and reduce systemic inflammation and disease severity.

The cholinergic system and Parkinson disease.

Although Parkinson disease (PD) is viewed traditionally as a motor syndrome secondary to nigrostriatal dopaminergic denervation, recent studies emphasize non-motor features. Non-motor comorbidities, such as cognitive impairment, are likely the result of an intricate interplay of multi-system degenerations and neurotransmitter deficiencies extending beyond the loss of dopaminergic nigral neurons. The pathological hallmark of parkinsonian dementia is the presence of extra-nigral Lewy bodies that can be accompanied by other pathologies, such as senile plaques. Lewy first identified the eponymous Lewy body in neurons of the nucleus basalis of Meynert (nbM), the source of cholinergic innervation of the cerebral cortex. Although cholinergic denervation is recognized as a pathological hallmark of Alzheimer disease (AD), in vivo neuroimaging studies reveal loss of cerebral cholinergic markers in parkinsonian dementia similar to or more severe than in prototypical AD. Imaging studies agree with post-mortem evidence suggesting that basal forebrain cholinergic system degeneration appears early in PD and worsens coincident with the appearance of dementia. Early cholinergic denervation in PD without dementia appears to be heterogeneous and may make specific contributions to the PD clinical phenotype. Apart from well-known cognitive and behavioral deficits, central, in particular limbic, cholinergic denervation may be associated with progressive deficits of odor identification in PD. Recent evidence indicates also that subcortical cholinergic denervation, probably due to degeneration of brainstem pedunculopontine nucleus neurons, may relate to the presence of dopamine non-responsive gait and balance impairments, including falls, in PD.

Age-associated leukoaraiosis and cortical cholinergic deafferentation.

OBJECTIVE: To investigate the relationship between age-associated MRI leukoaraiosis or white matter hyperintensities (WMH) and cortical acetylcholinesterase (AChE) activity. BACKGROUND: One possible mechanism of cognitive decline in elderly individuals with leukoaraiosis is disruption of cholinergic fibers by strategically located white matter lesions. Periventricular lesions may have a higher chance of disrupting cholinergic projections compared with more superficial nonperiventricular white matter lesions because of anatomic proximity to the major cholinergic axonal projection bundles that originate from the basal forebrain. METHODS: Community-dwelling, middle-aged and elderly subjects without dementia (mean age 71.0 +/- 9.2 years; 55-84 years; n = 18) underwent brain MRI and AChE PET imaging. The severity of periventricular and nonperiventricular WMH on fluid-attenuated inversion recovery MRI images was scored using the semiquantitative rating scale of Scheltens et al. [11C]methyl-4-piperidinyl propionate AChE PET imaging was used to assess cortical AChE activity. Age-corrected Spearman partial rank correlation coefficients were calculated. RESULTS: The severity of periventricular (R = -0.52, p = 0.04) but not nonperiventricular (R = -0.20, not significant) WMH was inversely related to global cortical AChE activity. Regional cortical cholinergic effects of periventricular WMH were most significant for the occipital lobe (R = -0.58, p = 0.02). CONCLUSIONS: The presence of periventricular but not nonperiventricular white matter hyperintensities (WMH) is significantly associated with lower cortical cholinergic activity. These findings support a regionally specific disruption of cholinergic projection fibers by WMH.

In vivo study of acetylcholine esterase in basal forebrain, amygdala, and cortex in mild to moderate Alzheimer disease.

It is currently unclear whether impairment of the cholinergic system is present in Alzheimer disease (AD) already at an early stage and to what extent it depends on degeneration of the nucleus basalis of Meynert (nbM). We examined acetylcholine esterase activity in vivo in the nbM, the amygdala, and cerebral neocortex. Measurements were performed in normal controls and in patients with mild to moderate AD with positron emission tomography (PET) and C-11-labeled N-methyl-4-piperidyl-acetate (MP4A) which is a specific substrate of AChE. AChE activity was reduced significantly in amygdala and cerebral cortex. In contrast, AChE activity and glucose metabolism appeared preserved or even increased in the nbM. The results support the concept that neocortical and amygdaloid functional changes of the cholinergic system are an early and leading event in AD, rather than the consequence of neurodegeneration of basal nuclei.

The distribution and morphological characteristics of catecholaminergic cells in the brain of monotremes as revealed by tyrosine hydroxylase immunohistochemistry.

The present study describes the distribution and cellular morphology of catecholaminergic neurons in the CNS of two species of monotreme, the platypus (Ornithorhynchus anatinus) and the short-beaked echidna (Tachyglossus aculeatus). Tyrosine hydroxylase immunohistochemistry was used to visualize these neurons. The standard A1-A17, C1-C3 nomenclature was used for expediency, but the neuroanatomical names of the various nuclei have also been given. Monotremes exhibit catecholaminergic neurons in the diencephalon (A11, A12, A13, A14, A15), midbrain (A8, A9, A10), rostral rhombencephalon (A5, A6, A7), and medulla (A1, A2, C1, C2). The subdivisions of these neurons are in general agreement with those of other mammals, and indeed other amniotes. Apart from minor differences, those being a lack of A4, A3, and C3 groups, the catecholaminergic system of monotremes is very similar to that of other mammals. Catecholaminergic neurons outside these nuclei, such as those reported for other mammals, were not numerous with occasional cells observed in the striatum. It seems unlikely that differences in the sleep phenomenology of monotremes, as compared to other mammals, can be explained by these differences. The similarity of this system across mammalian and amniote species underlines the evolutionary conservatism of the catecholaminergic system.

The distribution and morphological characteristics of cholinergic cells in the brain of monotremes as revealed by ChAT immunohistochemistry.

The present study employs choline acetyltransferase (ChAT) immunohistochemistry to identify the cholinergic neuronal population in the central nervous system of the monotremes. Two of the three extant species of monotreme were studied: the platypus (Ornithorhynchus anatinus) and the short-beaked echidna (Tachyglossus aculeatus). The distribution of cholinergic cells in the brain of these two species was virtually identical. Distinct groups of cholinergic cells were observed in the striatum, basal forebrain, habenula, pontomesencephalon, cranial nerve motor nuclei, and spinal cord. In contrast to other tetrapods studied with this technique, we failed to find evidence for cholinergic cells in the hypothalamus, the parabigeminal nucleus (or nucleus isthmus), or the cerebral cortex. The lack of hypothalamic cholinergic neurons creates a hiatus in the continuous antero-posterior aggregation of cholinergic neurons seen in other tetrapods. This hiatus might be functionally related to the phenomenology of monotreme sleep and to the ontogeny of sleep in mammals, as juvenile placental mammals exhibit a similar combination of sleep elements to that found in adult monotremes.

Morphine-induced spinal cholinergic activation: in vivo imaging with positron emission tomography.

Positron emission tomography (PET) imaging of spinal cord in monkeys with a cholinergic tracer demonstrates increased spinal cholinergic activity in response to an analgesic dose of morphine, and this PET result correlates with measurement of acetylcholine spillover into spinal cord extracellular space induced by morphine, as measured by microdialysis. Previous studies in rats, mice, and sheep demonstrate activation of spinal cholinergic neurons by systemic opioid administration, and participation of this cholinergic activity in opioid-induced analgesia. Testing the relevance of this observation in humans has been limited to measurement of acetylcholine spillover into lumbar cerebrospinal fluid. The purpose of this study was to apply a recently developed method to image spinal cholinergic terminals non-invasively via PET and to test the hypothesis that the tracer utilized would reflect changes in local cholinergic activity. Following Animal Care and Use Committee approval, seven adult male rhesus monkeys were anesthetized on three separate occasions. On two of the occasions PET scans were performed using [(18)F] (+)-4-fluorobenzyltrozamicol ([(18)F]FBT), which selectively binds to the vesicular acetylcholine (ACh) transporter in the presynaptic cholinergic terminals. PET scans were preceded by injection of either saline or an analgesic dose of IV morphine (10 mg/kg). On the third occasion, microdialysis catheters were inserted in the spinal cord dorsal horn and acetylcholine concentrations in dialysates determined before and after IV morphine injection. Morphine increased cholinergic activity in the spinal cord, as determined by blood flow corrected distribution volume of [(18)F]FBT in the cervical cord compared to the cerebellum. Morphine also increased acetylcholine concentrations in microdialysates from the cervical cord dorsal horn. The one animal which did not show increased spinal cholinergic activity by PET from this dose of morphine also did not show increased acetylcholine from this morphine dose in the microdialysis experiment. These data confirm the ability to use PET to image spinal cholinergic terminals in the monkey spinal cord and suggest that acute changes in cholinergic activity can be imaged with this non-invasive technique. Following preclinical screening, PET scanning with [(18)F]FBT may be useful to investigate mechanisms of analgesic action in normal humans and in those with pain.

Progressive loss of cortical acetylcholinesterase activity in association with cognitive decline in Alzheimer's disease: a positron emission tomography study.

We measured brain acetylcholinesterase activity in 30 patients with Alzheimer's disease (AD) and 14 age-matched controls by positron emission tomography (PET) and using a carbon 11-labeled acetylcholine analogue. Seven AD patients had repeat PET scans. The k3 values were calculated as an index of acetylcholinesterase activity in a three-compartment analysis using the metabolite corrected arterial input function. Twenty-eight of the 30 AD patients (14 each in the early and late onset subgroups) were retained in the study so as to equalize the range and average severity of cognitive impairment within the early and late onset subgroups. The k3 values were significantly reduced in the neocortex, hippocampus, and amygdala in the early onset AD patients, although the k3 values were significantly reduced only in the temporoparietal cortex and amygdala in the late onset AD patients. In the longitudinal study, all 7 repeat AD patients showed further reduction of cortical k3 values in the second PET scans, with a mean interval of 2 years, suggesting a progressive loss of the ascending cholinergic system from the nucleus basalis of Meynert in AD. In 37 AD patients, there was a highly significant correlation between the cortical k3 values and Mini-Mental State Examination scores, supporting the cholinergic hypothesis in AD.

Reproducibility of repeated measures of cholinergic terminal density using.

UNLABELLED: [18F](+)-4-fluorobenzyltrozamicol (FBT), which selectively binds to the vesicular acetylcholine transporter in the presynaptic cholinergic neuron, has previously been shown to be a useful ligand for the study of cholinergic terminal density in the basal ganglia with PET. The goal of this study was to assess the test-retest variability of [18F]FBT and PET measurements under baseline conditions in the basal ganglia. METHODS: After approval from the Animal Care and Use Committee, 6 rhesus monkeys underwent a series of 2 [18F]FBT PET scans (time between scans, 32-301 d) under isoflurane anesthesia. Each scan was initiated on the bolus injection of the radiotracer and consisted of 26 frames acquired during 180 min. Arterial blood samples were collected over the course of each scan to determine the metabolite-corrected arterial input function. Tissue time-activity curves were obtained from the scan data by drawing regions of interest over the basal ganglia and cerebellum. The distribution volume ratio for the basal ganglia was then determined for each scan by taking the ratio of the basal ganglia (specific binding) to cerebellum (nonspecific binding) distribution volume. Distribution volumes were derived using the Logan graphic analysis technique as well as a standard 3-compartment model. Additionally, the radioactivity concentration ratio was calculated as the ratio of the average [18F]FBT concentration in the basal ganglia to that in the cerebellum during the last half of the study (85-170 min). The constant K1, determined using the standard 3-compartment model, was used as an index of blood flow changes between studies. RESULTS: For all subjects, the test-retest variability was less than 15% for the distribution volume ratio and 12% for the radioactivity concentration ratio. Good agreement was found between the distribution volume ratio calculated using the graphic technique and the standard 3-compartment model. Using K1 as an index, the variability in blood flow seen in both the basal ganglia and the cerebellum was significantly reduced in their ratio. CONCLUSION: These results show the reproducibility of [18F]FBT and PET measurements in the basal ganglia.

In vivo imaging of the spinal cord cholinergic system with PET.

PURPOSE: Our goal was to demonstrate the feasibility of an in vivo noninvasive method for imaging spinal cord cholinergic terminals using (+)-4-[18F]fluorobenzyltrozamicol ([18F]FBT) and PET. METHOD: In vitro and in vivo experiments in rats were conducted to demonstrate the specific binding characteristics, localization, and time course of [3H]FBT binding in the spinal cord. PET imaging was then performed on seven rhesus monkeys. RESULTS: The rat studies demonstrate high specific binding in the spinal cord with a distribution coinciding with the known distribution of cholinergic terminals. In vivo tracer concentrations in the spinal cord and basal ganglia were of the same magnitude. With use of [18F]FBT and PET in the rhesus monkey, the spinal cord was clearly visualized, with tracer concentration in the spinal cord being approximately one-fourth of that seen in the basal ganglia. CONCLUSION: This work demonstrates the feasibility of imaging cholinergic terminals in vivo in the spinal cord using [18F]FBT and PET.

Muscarinic receptor loss and preservation of presynaptic cholinergic terminals in hippocampal sclerosis.

PURPOSE: Prior single-photon emission tomography studies showed losses of muscarinic acetylcholine receptor (MAChR) binding in patients with refractory mesial temporal lobe epilepsy. Experimental animal studies demonstrated transient losses of MAChR due to electrically induced seizures originating in the amygdala. However, the relations between cholinergic synaptic markers, seizures, and underlying neuropathology in human temporal lobe epilepsy are unknown. We tested the hypotheses that human brain MAChR changes are attributable to hippocampal sclerosis (HS), and that HS resembles axon-sparing lesions in experimental animal models. METHODS: We measured MAChR binding-site density, an intrinsic neuronal marker, within the hippocampal formation (HF) in anterior temporal lobectomy specimens from 10 patients with HS and in 10 autopsy controls. Binding-site density of the presynaptic vesicular acetylcholine transporter (VAChT) was measured as a marker of extrinsic cholinergic afferent integrity. MAChR and VAChT results were compared with neuronal cell counts to assess their relations to local neuronal losses. RESULTS: Reduced MAChR binding-site density was demonstrated throughout the HF in the epilepsy specimens compared with autopsy controls and correlated in severity with reductions in cell counts in several HF regions. In contrast to MAChR, VAChT binding-site density was unchanged in the epilepsy specimens compared with autopsy controls. CONCLUSIONS: Reduction in MAChR binding in HS is attributable to intrinsic neuronal losses. Sparing of afferent septal cholinergic terminals is consistent with the hypothesis that an excitotoxic mechanism may contribute to the development of HS and refractory partial epilepsy in humans.

Brain muscarinic receptors in progressive supranuclear palsy and Parkinson's disease: a positron emission tomographic study.

OBJECTIVES: To assess muscarinic acetylcholine receptors (mAChRs) in the brains of patients with progressive supranuclear palsy and Parkinson's disease, and to correlate the cholinergic system with cognitive function in progressive supranuclear palsy and Parkinson's disease. METHODS: Positron emission tomography (PET) and [11C]N-methyl-4-piperidyl benzilate ([11C]NMPB) was used to measure mAChRs in the brain of seven patients with progressive supranuclear palsy, 12 patients with Parkinson's disease, and eight healthy controls. All of the patients with progressive supranuclear palsy were demented. The Parkinson's disease group consisted of 11 non-demented patients and one demented patient. The mini mental state examination (MMSE) was used to assess the severity of cognitive dysfunction in all of the subjects. The modified Wisconsin card sorting test (WCST) was used to evaluate frontal cognitive function in the non-demented patients with Parkinson's disease and controls. RESULTS: The mean K3 value, an index of mAChR binding, was significantly higher for the frontal cortex in the patients with Parkinson's disease than in the controls (p<0.01). By contrast, the patients with progressive supranuclear palsy had no significant changes in the K3 values of any cerebral cortical regions. The mean score of the MMSE in the progressive supranuclear palsy group was significantly lower than that in the control group. Although there was no difference between the Parkinson's disease and control groups in the MMSE, the non-demented patients with Parkinson's disease showed significant frontal lobe dysfunction in the WCST. CONCLUSIONS: The increased mAChR binding in the frontal cortex of the patients with Parkinson's disease may reflect denervation hypersensitivity caused by loss of the ascending cholinergic input to that region from the basal forebrain and may be related to frontal lobe dysfunction in Parkinson's disease. The cerebral cortical cholinergic system may not have a major role in cognitive dysfunction in progressive supranuclear palsy.

Distribution of choline acetyltransferase (ChAT) immunoreactive nerve fibers in the nasal mucosa; especially in the respiratory epithelium.

The distribution and morphological construction of cholinergic nerve fibers in the respiratory nasal mucosa of the rat and human were investigated using choline acetyltransferase (ChAT) activities by means of light microscopy, confocal laser scanning microscopy and electron microscopy. It was observed that ChAT-immunoreactive (IR) nerve fibers were distributed to form fine varicosities around the blood vessels and seromucous glands under the epithelium and at the basement membrane and within the epithelium. Electron microscopy showed that the ChAT-IR nerve fibers within the epithelium terminated as free nerve endings. Some of them were in close contact with goblet cells. The distribution of ChAT immunoreactive fibers resembled that of vasoactive intestinal polypeptide-IR nerve fibers.

Radioiodinated 2-hydroxy-3-(4-iodophenyl)-1-(4-phenylpiperidinyl)propane: potential radiotracer for mapping central cholinergic innervation in vitro.

Radioiodinated 2-hydroxy-3-(4-iodophenyl)-1-(4-phenylpiperidinyl)propane, 5 (4-HIPP), was synthesized and evaluated as a simple vesamicol-like radiotracer for mapping cholinergic pathways in the brain. Both enantiomers of 5 exhibit significant accumulation (approx. 2% of injected dose) and prolonged retention (t1/2 greater than 3 h) within the rat brain. The accumulation of radioiodinated 5 in the rat brain was reduced by up to 70% in the presence of vesamicol and its analogs. The levorotary isomer (-)-4-[123I]HIPP exhibits significant accumulation in the monkey brain, with a half-life of about 9 h. Radioiodinated 5 may therefore be a useful tool for studying cholinergic pathways in the brain.

Positron emission tomographic studies of central cholinergic nerve terminals.

The aim of this study was to develop a quantitative method for the study of cholinergic nerve terminals in vivo. An 18F-labeled analogue of vesamicol ([18F]FMV) that binds with high affinity to synaptic vesicles from Torpedo electric organ was synthesized and evaluated in vivo in rats and monkeys by positron emission tomography (PET). In rats, the tracer was rapidly cleared from the blood and highly extracted into the brain, where it was specifically and irreversibly bound. In monkeys, a specific binding of the tracer was observed in brain regions known to contain cholinergic nerve terminals. Preinjection of non-labeled vesamicol prevented the cerebral binding of [18F]FMV to a high affinity site in both species. Our results are a major step towards quantitative human in vivo studies of presynaptic cholinergic functions.