Distribution Of Intracellular Receptors Research Articles

Differential Distribution of Intracellular Glutamate Receptors in Dendrites

Glutamate receptors are synthesized in the cell body and transported in intracellular compartments to the target synapse. The objective of the present study was to analyze the intracellular pool of glutamate receptors and determine whether the intracellular pool was related to the synaptic distribution of the receptors. As a model system, we chose the fusiform cell of the dorsal cochlear nucleus for which we have previously demonstrated that receptors are selectively targeted to synapses on apical and basal dendrites. A combination of retrograde tracing and postembedding immunogold labeling was used to quantify intracellular receptors in segments of apical and basal dendrites. Immunolabeling for GluR4 and mGluR1alpha is present at synapses on basal dendrites but not on apical dendrites, whereas immunolabeling for GluR2/3 is present at both populations of synapses. In the analysis of intracellular pools, we find that GluR2/3 is equally distributed in apical and basal dendrites, whereas GluR4 and mGluR1alpha are more concentrated in basal dendrites than in apical dendrites. These findings indicate that the distribution of intracellular receptors is related to that of synaptic receptors and suggest that a mechanism exists in neurons to target proteins to dendritic domains soon after synthesis. We found no evidence for the existence of a pool of intracellular receptors, which could represent a receptor reserve, near the postsynaptic density. Receptors were often found in clusters associated with tubulovesicular membranes of the endoplasmic reticulum, identified with immunoglobulin binding protein (BIP) or calnexin, suggesting that this organelle is involved in receptor transport in dendrites.

The glucocorticoid receptor in GH1 cells. Evidence from dense amino acid labeling and whole cell studies for an equilibrium model explaining the influence of hormone on the intracellular distribution of receptor.

The glucocorticoid receptor in GH1 cells cultured without glucocorticoids and harvested in hypotonic buffer is primarily a cytosolic protein with a sedimentation coefficient of 10 S in low salt gradients. In gradients containing 0.4 M KCl, the receptor sediments at 3.5 S. Following incubation of cells for various times in culture medium containing dense amino acids, discrete peaks of newly synthesized dense receptor and normal density 3.5 S receptor are resolved in salt-containing gradients. The half-life of the 3.5 S receptor is 9.5 h. In contrast, the 10 S receptor gradually becomes more dense as the time of incubation of cells with dense medium increases and discrete normal and dense receptor peaks are not resolved in sucrose gradients. These results suggest that the 10 S receptor is an oligomer and is in a rapidly exchanging equilibrium in intact cells at 37 degrees C with its 3.5 S hormone-binding subunit. When GH1 cells are incubated for 1 h at 37 degrees C with various concentrations of [3H]9 alpha-fluoro-11 beta, 16 alpha, 17 alpha, 21-tetrahydroxypregna-1,4-diene-3,20-dione, 16,17-acetonide ([3H]triamcinolone acetonide), receptor is found both in the cytosol and the nucleus. Sedimentation of the cytosol in low salt gradients reveals that a 4 S receptor form is present along with 10 S receptor. As the concentration of [3H]triamcinolone acetonide is increased, the amounts of both 4 S cytosolic receptor and nuclear receptor increase and the amount of 10 S cytosolic receptor decreases. Following removal of [3H]triamcinolone acetonide from cells, the amounts of both 4 S cytosolic receptor and nuclear receptor decrease and the amount of 10 S cytosolic receptor increases. Density labeling indicates that this increase in 10 S receptor is not due to new receptor synthesis. The 4 S receptor form, but not the 10 S form, binds to purified DNA. Resedimentation studies suggest that the 4 S receptor found in low salt gradients is identical with or derived from the 3.5 S form found in salt-containing gradients. A model for hormone-mediated receptor translocation to the nucleus is proposed in which binding of hormone shifts the equilibrium between the oligomeric 10 S receptor and its 4 S subunit toward the 4 S form. The 4 S form is in equilibrium with 4 S receptor bound to chromatin. The model provides a molecular explanation for many experimental findings concerning glucocorticoid and other steroid receptors.

A direct effect of LHRH on anterior pituitary estrogen receptors in the female rat

An analysis has been made of the effects of LHRH on anterior pituitary estrogen receptors. Dispersed pituitary cells from ovariectomized, estrogen-primed rats were incubated in the presence or absence of 1–10 ng of LHRH per pituitary equivalent for 30 min at 37°C. Following lysis, nuclear estrogen receptor concentration was observed to be significantly elevated in a dose-related fashion, while extranuclear binding was depressed by a constant factor. At 5 or 10 ng LHRH levels, the increased nuclear binding could not be explained by translocation of cytoplasmic receptor, and total cellular receptor activity was significantly enhanced. Isolated cytosol and nuclei were not subject to significant fluctuations in the presence of 1–10 ng of LHRH, although 100 or 1000 ng of LHRH did significantly stimulate nuclear binding. In separate experiments, 15 min incubations with 1 or 10 ng of LHRH were not sufficient to cause altered receptor binding, under conditions where 30 min incubation effected the above results. At 120 min of incubation, binding in the extranuclear compartment was increased dramatically over the 30-min interval, and this was entirely at the expense of the nuclear binding capacity. In vivo administration of LHRH caused an increase in nuclear anterior pituitary receptor activity at 1 h, without concomitant alteration in cytosol content. At 10 h, the intracellular distribution of receptors had shifted in a manner consistent with nuclear→cytoplasmic recycling of receptor, and the distribution mirrored that of the untreated control animals. The results indicate that LHRH has a direct stimulatory effect on nuclear estrogen receptor activity which cannot be explained by stimulation of cytoplasmic receptor depletion. Such an action could account for the self-priming effect of LHRH on its own activity, since estrogens are known to modulate (presumably through a receptor-mediated mechanism of action) the responsiveness of pituitary cells to LHRH.