Variations In Receptor Density Research Articles

Density of available striatal dopamine receptors predicts trait impulsiveness during performance of an attention-demanding task.

The density (measured at binding potential) of available striatal D2/D3 receptors has been shown to predict trait impulsiveness. This relationship is highly robust and well replicated. In each case, however, the availability of dopamine receptors was measured at rest. More broadly, the extent to which relationships between dopamine receptor availability and behavioral traits hold when participants perform a cognitive task is unclear. Furthermore, the performance of a cognitive task engages fundamentally different neural networks than are maximally engaged during the resting state. This complicates interpretation of previously observed correlations, which could be influenced by two distinct factors. The first is variation in available receptor density, which reflects a stable trait of the individual. The second is variation in context-specific dopamine release, which differentially displaces some dopamine radiotracers (such as raclopride) across individuals. Using an existing data set, we related trait impulsiveness, as measured using the Barratt Impulsiveness Scale (BIS-11), to the density (binding potential) of available striatal D2/D3 receptors as measured using positron emission tomography (PET) with [11C]raclopride. Importantly, the PET scan was completed while participants performed an attention-demanding visual search task. We replicate robust correlations between this measure of receptor availability and trait impulsiveness previously demonstrated during the resting state, extending this relationship to periods of active task engagement. Our results support the idea that this relationship depends on striatal D2/D3 receptor density and not on context-dependent dopamine release.NEW & NOTEWORTHY Several studies have demonstrated a relationship between the density of available striatal D2/D3 receptors and trait impulsiveness. However, in each case, the availability of dopamine receptors was measured during the resting state. This complicates interpretation of previously observed correlations, which could be influenced by either stable variation in receptor density or context-dependent dopamine release. We present evidence uniquely consistent with the former interpretation, providing clarity to the nature of this brain-behavior relationship.

A single-scan protocol for absolute D2/3 receptor quantification with [123I]IBZM SPECT

PurposeMolecular imaging of the D2/3 receptor is widely used in neuropsychiatric research. Non-displaceable binding potential (BPND) is a very popular quantitative index, defined as the product of the receptor concentration (Bavail) and the radiotracer affinity for the receptor (1/appKd). As the appKd is influenced by parameters such as the endogenous neurotransmitter dynamics, it often constitutes a confounding factor in research studies. A simplified method for absolute quantification of both these parameters would be of great interest in this context. Here, we describe the use of a partial saturation protocol that permits to produce an in vivo Scatchard plot and thus estimate Bavail and appKd separately, through a single dynamic SPECT session. To validate this approach, a multi-injection protocol is used for the full kinetic modeling of [123I]IBZM using a 3-tissue compartment, 7-parameter model (3T-7k). Finally, more “classic” BPND estimation methods are also validated against the results of the 3T-7k. MethodsTwenty-nine male rats were used. Binding parameters were estimated using the 3T-7k in a multi-injection protocol. A partial saturation protocol was applied at the region- and voxel-level and results were compared to those obtained with the 3T-7k model. The partial saturation protocol was applied after an adenovirus-mediated D2 receptor striatal overexpression and in an amphetamine-induced dopamine release paradigm. The Simplified Reference Tissue Model (SRTM), the Logan's non-invasive graphical analysis (LNIGA) and a simple standardized uptake ratio (SUR) method were equally applied. ResultsThe partial saturation experiments gave similar values as the 3T-7k both at the regional and voxel-level. After adenoviral-mediated D2-receptor overexpression, an increase in Bavail by approximately 18% was observed in the striatum. After amphetamine administration, a 16.93% decrease in Bavail (p<0.05) and a 39.12% increase (p<0.01) in appKd was observed. BPND derived from SRTM, LNIGA and SUR correlated well with the Bavail values from the 3T-7k (r=0.84, r=0.84 and r=0.83, respectively, p<0.0001 for all correlations). ConclusionA partial saturation protocol permits the non-invasive and time-efficient estimation of Bavail and appKd separately. Given the different biological phenomena that underlie these parameters, this method may be applied for the in-depth study of the dopaminergic system in translational molecular imaging studies. It can detect the biological variations in these parameters, dissociating the variations in receptor density (Bavail) from affinity (1/appKd), which reflects the interactions of the receptor with its endogenous ligand.

Location, location, location: Variation in sensitivity to pain across the body.

There is evidence that sensitivity to noxious stimuli differs between the sexes and across the body, but few studies have investigated differences in the perception and experience of acute pain stimuli across the body in healthy individuals. We recruited 52 healthy participants, aged 18-36 (50% men) and administered 39, 42 and 45°C stimuli at four body sites bilaterally to examine differences in the experience of pain intensity and unpleasantness between body sites via an 11-point numerical rating scale. Noxious and innocuous thermal heat stimuli were perceived as significantly more intense when delivered to the wrist (M=3.98, SD = 1.93) and back (M =4.07, SD=1.98) compared to the shoulder (M=3.45, SD=1.91) and leg (M=3.46, SD=1.87). Pain unpleasantness ratings yielded similar findings; stimuli were perceived as more unpleasant when administered to the wrist (M=2.83, SD=1.93) and lower back (M=3.04, SD=2.11) compared to the shoulder (M=2.63, SD=1.85) and leg (M=2.26, SD=1.82). These findings suggest that painful thermal stimuli delivered to the wrist and back are perceived as more intense and unpleasant compared with other body sites in healthy persons. These differences may be due to variations in receptor density, or the relative importance of these sites for daily living and survival. Moreover, these insights are helpful for the design of studies investigating pain experience in healthy persons in experimental or clinical settings. WHAT DOES THIS STUDY ADD?: We tested sensitivity to acute suprathreshold thermal stimulations across a range of body sites to investigate for potential variability. We found significant differences in the perceived intensity and unpleasantness of noxious and innocuous thermal stimuli at the wrist and lower back, compared with the shoulder and leg. These results suggest that pain experience is driven by receptor density or the relative functional importance of these sites.

Unraveling the Binding Mechanism of Trivalent Tumor Necrosis Factor Ligands and Their Receptors

Characterization of the binding of a tumor necrosis factor (TNF) ligand to its receptor(s) is pivotal to understand how these proteins initiate signal transduction pathways. Unfortunately, kinetic elucidation of these interactions is strongly hampered by the multivalent nature of the binding partners. The interaction between TNF-related apoptosis-inducing ligand and its death receptors was analyzed using in-depth applications of surface plasmon resonance technology. Variations in receptor density and sensor chip type allowed us to manipulate the stoichiometry of the formed complex, and the rate constants describing the binding of trimeric TNF-related apoptosis-inducing ligand to only one receptor molecule were determined. Remarkably, the affinity of this trimer-monomer complex is in the picomolar range, and its dissociation very slow. Further analysis showed that the second and third receptor molecules bind with lower affinity to the preformed trimer-monomer complex. This together with results obtained with receptor activator of NF-κB ligand and B cell-activating factor strongly suggests that the binding of TNF family ligands to their receptors is initiated via the formation of a trimer-monomer complex that is sufficiently stable to allow binding of two additional receptor molecules. These results suggest that avidity does not play a significant role and thus provide new insight in how TNF ligands form the biologically important complexes with their receptors.

Autoradiographic analysis of ghrelin receptors in the rat hypothalamus

Ghrelin exerts potent stimulatory effects on food intake. It is assumed to increase feeding by binding at growth hormone secretagogue receptors (GHS-R), the only sites of action for this gastric hormone identified to date. Initially, the distribution of ghrelin binding sites could only be determined from expression patterns of GHS-R mRNA or the use of immunohistochemical techniques to examine c-fos expression. However, the characterisation of a novel radioligand ([ 125I- his 9]-ghrelin), has enabled the distribution of GHS-R receptor protein to be directly demonstrated. Here, using quantitative autoradiography, we investigate the distribution and density of ghrelin receptors in the rodent hypothalamus. Specific binding was identified in the appetite-regulating arcuate nucleus, ventromedial hypothalamic nucleus, paraventricular nucleus, dorsomedial hypothalamic nucleus and the lateral hypothalamic area corresponding to the previously reported distribution pattern of GHS-R mRNA. Surprisingly, variations in receptor density were not identified in any of these binding sites upon a change in nutritional status, despite relevant alterations in plasma ghrelin levels being identified. We suggest that this may relate to the paradigm employed to modify nutritional status in the study or could indicate that peripheral ghrelin is unlikely to be the major source of ghrelin that acts in many hypothalamic sites.

Vibrotactile thresholds at the fingertip, volar forearm, large toe, and heel

Thresholds for the perception of vibration vary with location on the body due to the organization of tactile channels in hairy and non-hairy skin, and variations in receptor density. This study determined vibration thresholds at four locations on the body with two different contactors so as to assist the identification of the tactile channel determining the threshold at each location. Vibrotactile thresholds at six frequencies from 8 to 250 Hz were measured on the distal phalanx of the index finger, the volar forearm, the large toe, and the heel with two contactors: (i) a 1-mm diameter circular probe with a 1-mm gap to a fixed circular surround (i.e., 7.1-mm2 excitation area), and (ii) a 6-mm diameter circular probe with a 2-mm gap to a fixed circular surround (i.e., 79-mm2 excitation area). At all frequencies and with both contactors, thresholds on the fingertip were lower than thresholds on the volar forearm, the large toe, and the heel, consistent with a greater density of mechanoreceptors at the fingertip. Thresholds with the larger contactor were lower than thresholds with the smaller contactor on the fingertip at high frequencies (63, 125, and 250 Hz), on the large toe (except at 250 Hz), on the heel (at all frequencies), and on the volar forearm at 250 Hz. It is concluded that at least two tactile channels (Pacinian from 63 to 250 Hz, and non-Pacinian from 8 to 31.5 Hz) determined vibrotactile thresholds at the fingertip, whereas non-Pacinian channels had a dominant influence on vibrotactile thresholds at the volar forearm. The role of Pacinian and non-Pacinian channels could not be confirmed at the large toe or the heel despite some evidence of spatial summation.

Platelet PAR1 receptor density—Correlation to platelet activation response and changes in exposure after platelet activation

IntroductionA polymorphism (-14 A/T) affecting PAR1 expression on the platelet surface has recently been identified. A two-fold variation in receptor density, which correlated with the platelet response to PAR1-activating peptide (PAR1-AP), has been reported. Materials and methodsWe used flow cytometry to measure the correlation between the number of PAR1 receptors and platelet activation. We also measured the changes in receptor exposure after platelet activation with PAR1-AP, ADP, PAR4-AP or a collagen-related peptide (CRP). ResultsIn our study, the PAR1 receptor number varied almost four-fold, from 547 to 2063 copies/platelet (mean±S.D. 1276±320, n=70). The number of PAR1 receptors on resting platelets correlated to platelet fibrinogen binding and P-selectin expression following platelet activation with PAR1-AP (r2=0.30, p<0.01 and r2=0.15, p<0.05, respectively, n=36). The correlation was not improved by exclusion of the ADP-component from the PAR1-AP-induced response. We found a trend, but no statistically significant differences in PAR1 receptor number and platelet reactivity between A/A individuals and T/A or T/T individuals. Ex vivo activation with PAR1-AP decreased PAR1 surface exposure to 71±19% of the exposure on resting platelets (mean±S.D., p<0.01, n=19), while activation by ADP, PAR4-AP or CRP significantly increased the exposure, to 151±27%, 120±21% and 138±25%, respectively (n=11, 11 and 10). ConclusionsThis study shows a large variation in PAR1 receptor number in healthy individuals, a variation correlated to the platelet activation response. We found a significant reduction in PAR1 surface exposure after adding PAR1-AP, while activation with ADP, PAR4-AP or CRP increased the exposure.

Hereditary Variation in Platelet Integrin α2β1 Density Is Associated With Two Silent Polymorphisms in the α2 Gene Coding Sequence

AbstractThe integrin α2β1 is a receptor for collagen that plays a fundamental role in the adhesion of blood platelets to the extracellular matrix. We previously reported that platelet α2β1 levels among randomly selected individuals can vary up to 10-fold and that this correlates with differences in adhesiveness to type-I or type-III collagens. We have now found two linked, allelic polymorphisms within the coding sequence of the α2 gene that correlate with receptor density, TTT/TTC at codon Phe224 and ACA/ACG at codon Thr246. By Southern blot hybridization of specific antisense DNA probes to segments of genomic DNA that encompass each coding region, we have determined the gene frequencies of each allele in a random donor population (n = 65) to be 0.585 (TTC ... ACG) and 0.415 (TTT ... ACA). There is a statistically significant correlation between the alleles TTT ... ACA (codons 224…246) and high receptor density (n = 30; P < .002), whereas the complimentary alleles TTC ... ACG are associated with low receptor density. Heterozygous individuals express intermediate levels of this receptor, and familial studies confirm that these allelic polymorphisms are inherited characteristics. These findings prove that the level of platelet α2β1 is an inherited trait. The molecular basis for receptor density remains to be determined, but our findings establish that these silent alleles within the coding sequence of the α2 gene are linked to the genetic basis for variation in receptor density.

Localization of nicotinic cholinergic receptors in rat brain: Autoradiographic studies with [ 3H]cytisine

There is a great deal of interest in the role of nicotinic acetylcholine receptors in the central nervous system, although their function is not well understood at present. Currently, central nicotinic receptors can be classified broadly as either α-bungarotoxin binding sites with low affinity for acetylcholine agonists, or as high-affinity agonist binding sites with low affinity for a-bungarotoxin. Neuronal nicotinic receptors with a high affinity for agonists are distributed widely in the central nervous system. Evidence from molecular biology and electrophysiology suggests that multiple nicotinic receptor types exist in the brain. In this study we have used the agonist [ 3H]cytisine as a ligand for autoradiography to generate a detailed quantitative map of the high-affinity agonist binding nicotinic receptor in the rat brain. Optimized binding conditions, characterization of the kinetic and equilibrium binding properties, and demonstration of the nicotinic pharmacology of this binding site in tissue sections confirm the usefulness of [ 3H]cytisine as a ligand for nicotinic receptor autoradiography. [ 3H]Cytisine autoradiography provides excellent anatomic resolution with very low non-specific binding. This property has allowed us to describe variations in receptor density within subnuclei and gradients of receptor density in larger brain regions. Data from several studies suggest that the predominant high-affinity agonist binding nicotinic receptor in the central nervous system is composed of the α4 and β2 subunits. The data in the current study are consistent with the suggestion that [ 3H]cytisine labels only the α4β2 nicotinic receptor with high affinity, offering the possibility of localizing a specific nicotinic receptor subtype in the central nervous system. In summary, we characterize the optimum experimental conditions for the use of [ 3H]cytisine in tissue section autoradiography. [ 3H]Cytisine proves to be an excellent marker for nicotinic cholinergic receptors with a very high affinity and very low background. We provide a detailed quantitative characterization of nicotinic receptor density in the rat central nervous system and we find there are significant variations and gradients in receptor density within specific brain regions, including subrogions previously thought to be homogeneous.

Differential distribution of excitatory amino acid receptors on embryonic rat spinal cord neurons in culture

1. Excitatory amino acid (EAA) receptors mediate synaptic transmission in many areas of the vertebrate CNS. To map the distribution of the EAA receptors, three agonists selective for EAA receptor subtypes [kainate, quisqualate, and N-methyl-D-aspartate (NMDA)] were applied by pressure to the cell bodies and neurites of voltage-clamped, embryonic rat spinal cord neurons in culture. 2. Current loss along the neurite between the site of activation and the recording electrode at the soma was estimated independently of variations in receptor density by focal application of high [K+] solution. This estimate was used to compensate agonist-evoked responses for current loss due to leak. K(+)-evoked current amplitudes progressively decreased as applications were made at more distal positions along the neurite. 3. Response amplitudes to EAAs showed a monotonic decay, similar to that with high [K+] solution, in only a few of the cells tested with each agonist. The majority of neurons had areas of high agonist sensitivity along the neurites, implying a nonuniform density of receptors. Such regions sometimes occurred at the most proximal segment of the neurite. Most neurites had distal regions of agonist sensitivity that ranged from 40 to 300% higher than at the soma. 4. The relative distributions of the three EAA receptors were compared by applying two agonists to the same sites along neurites and observing variations in normalized response amplitudes. When comparing NMDA versus non-NMDA receptor distributions on the same neurite, approximately 40% of the cells had similar patterns to two agonists, and the remaining 60% had different patterns. When sensitivity to the two non-NMDA agonists kainate and quisqualate were compared, about one-half of the cells tested had different patterns of agonist sensitivity. 5. Areas of high sensitivity persisted after block of calcium channels by addition of La3+ to the bath solution and after prevention of evoked transmitter release by a low [Ca2+]/high [Mg2+] solution. 6. These results show that spinal cord neurons can have regions of high agonist sensitivity to NMDA, kainate, and quisqualate along their neurites and that the sensitivity to any one of the EAA receptor subtypes can be elevated independently of the others.

Variation among acetylcholine receptor clusters induced by ciliary ganglion neurons in vitro

We have examined the variation in receptor density and area among neurite-associated acetylcholine receptor patches (NARPs) induced by chick ciliary ganglion neurons on nearby myotubes in vitro. Quantitative analysis of rhodamine-α-bungarotoxin (RBTX) NARPs revealed that about 15% of the NARPs were “outstanding” in terms of size (>60 μm 2) and fluorescence intensity (>100 units on a 0–255 scale). The total number of receptors at different NARPs ranged over 3 orders of magnitude. It is likely that variation in NARP size and intensity reflects regional variation in the ability of myotubes to respond to the neuronal influence because (1) no gradient in NARP size or intensity with distance from the soma was evident; (2) the intensities and areas of uninnervated receptor clusters (hot spots) were similar to those of NARPs; (3) acetylcholinesterase was present at the same proportion of hot spots and NARPs at all times examined. We found no physiological or morphological evidence that outstanding NARPs were more effective sites of transmitter release. Outstanding NARPs were restricted to the longest neurite of individual neurons, so they may signal trophic interactions of the sort that promote neurite outgrowth and survival.

Effect of vagotomy on cholinergic parameters in nuclei of rat medulla oblongata

Cholinergic enzymes and muscarinic receptors in nuclei of rat medulla oblongata were examined after unilateral vagotomy to determine their association with efferent vagal neurons. Vagotomy caused an ipsilateral depletion of acetylcholinesterase from the dorsal motor nucleus of the vagus (DNV) and the nucleus ambiguus (NA). Choline acetyltransferase activity was reduced in ipsilateral DNV, nucleus tractus solitarius and rostral NA. Muscarinic receptor localization by autoradiography with [ 3H]quinuclidinyl benzilate (QNB) revealed marked intranuclear variations in receptor density. Vagotomy had no effect on the QNB binding pattern. Loss of cholinergic enzymes is a consistent response of motor and preganglionic autonomic neurons to axotomy. Depletion of muscarinic receptors is an additional component of axon reaction in brain stem motoneurons. Accordingly, previous studies have shown a decrease in neurotransmitter-related proteins after axotomy of motoneurons. In the present study, cholinergic enzymes were depleted from axotomized vagal neurons but receptors were not. It is concluded that muscarinic receptors in the DNV and NA are not associated with vagal efferent neurons.