Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “In vitro model”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 595 records · Page 33Linked to original sources
PubMed2005

Characterization of the rotating dialysis cell as an in vitro model potentially useful for simulation of the pharmacokinetic fate of intra-articularly administered drugs.

The rotating dialysis cell consisting of a donor and an acceptor compartment with volumes of 10 and 1000 ml, respectively, separated by a dialysis membrane is proposed as an in vitro model potentially useful for simulation of the events influencing drug residence time in the knee joint cavity after intra-articular instillation. The purpose of this study was to characterize the rotating dialysis cell model with respect to basic model and drug related factors affecting the rate of drug appearance in the acceptor phase after initial instillation of the solutes into the donor cell. A total of 15 model compounds were included in the study and it was revealed that the transport processes were governed by (i) the volume of the donor solution and (ii) the molecular weight of the diffusants. A relationship between the donor volume-independent permeability coefficient and the molecular weight of the diffusants has been established. Additionally, the model was robust with release kinetics being insensitive to changes in pH, ionic strength, viscosity of the release medium, and revolution speed of the donor cell. The characteristics of the rotating dialysis cell model suggest that it may be a useful tool in the design of innovative depot injectables in the area of local joint delivery.

Dialysis↗
PubMed1997

Felodipine protects human atrial muscle from hypoxia-reoxygenation dysfunction: a force-frequency relationship study in an in vitro model of stunning.

AIMS: We aimed at investigating contractile changes after hypoxia-reoxygenation and dobutamine challenge in superfused human atrial pectinate muscle to see whether high versus low stimulation rate during hypoxia might account for outcome differences compatible with the definition of an in vitro model of myocardial stunning and whether pretreatment with the dihydropyridine Ca2+ entry blocker felodipine might afford protection. METHODS: Human right atrial trabeculae obtained from adult patients were superfused in an organ bath with oxygenated (O2 content 16 ml/l) and modified (NaHCO3 25.7 mmol/l) Tyrode's solution at 37 degrees C. Dobutamine (1 nmol/l to 10 micromol/l) was superfused in 10 oxygenated preparations to select the optimal drug concentration to be used in another 22 which were randomized. Group (A) consisted of time-related controls (Tyrodes's solution for 225 min at cycle length (CL) 1600 ms and no dobutamine). There were two test groups, respectively: (B) low (1600 ms CL) and (C) high (400 ms CL) stimulation rate. After 60 min of stabilization, in groups B and C, hypoxic superfusion (O2 content 5 ml/l) lasted 60 min, then reoxygenation (60 min) and dobutamine challenge (1 micromol/l, 15 min) were performed. Analysis of variance for repeated measures with the Greenhouse-Geisser correction, and a repeated measures model with structured covariance (preparation mass, length, width and time-varying time to peak tension) matrices were used whereby grouping (G), time (T) and G x T interaction were weighted. Force-frequency relationship and post-pausal potentiation were studied after each phase. Electrophysiology, histomorphometry and electron microscopy were carried out (n=6). Felodipine (0.1 micromol/l, n=5) pretreatment (15 min before hypoxia) was given in parallel experiments. RESULTS: Time-related controls showed approximately 10% per hour decrease of developed tension and the Paradise test provided approximately 80% of control values. In test groups (as compared to baseline values) contractility was decreased approximately 65% after hypoxia-reoxygenation and it increased approximately 25% after dobutamine (G, 0.0065<P<0.0155; T, P=0.00005; G x T, P=0.00005). High stimulation rate during hypoxia worsened hypoxia-reoxygenation contractile changes, whereas reversibility after dobutamine was less. In both B and C groups during hypoxia, contractility decreased quite rapidly, although by 10 min or so a plateau (approximately 50%) was reached in group B, whereas in group C contractility decreased to <20%. None of the covariates contributed significantly to predict the dependent variables investigated. Force-frequency relationship and post-pausal potentiation were repeatable, paralleled overall changes due to hypoxia, reoxygenation and dobutamine challenge and were useful to discriminate Ca2+-related diastolic processes thus helping index myocardial contractile reserve. Force-frequency relationship was negative at high stimulation rates, concomitant to an abrupt change of shape and duration of action potential with little time for Ca2+-related Ca2+ release and ensuing systolic processes. Felodipine pretreatment enabled an unblunted response to dobutamine. Histomorphometry showed an unexpected 'fibrotic core'. At electron microscopy, subendocardial and deep part of the same pectinate muscles showed identical degrees of degenerative lesions. Superfused samples showed, unexpectedly, less anoxic lesions than preparations fixed within 15 min from surgical explant, although lesions were higher than in samples fixed immediately after explant. CONCLUSIONS: This might be a relevant model, whereby pharmacological or physical interventions are tested. Native human atrial trabeculae might be used without dissection and/or preservatives. If high stimulation rate during hypoxia is used the power of hypothesis testing is maximized. Future studies with this material will be easier and comparatively smaller series might be investigated. Felo

Adult↗
PubMed1996

New pharmacokinetic in vitro model for studies of antibiotic activity against intracellular microorganisms.

The capacity for intracellular growth is an important survival strategy for a large group of common pathogens. Helicobacter pylori, the etiological agent for gastritis and duodenal ulcer, has been shown by both in vivo and in vitro studies to have the capacity to invade epithelial cells. In vitro models are used to study the effect of antibiotics on microoganisms. Most investigations are performed in broth culture or on agar plates, but kinetic models for bacteria in broth have been described. We present a new, kinetic model adapted for intracellular pathogens. A glass chamber, with a metal rack fitting Falcon cell culture inserts, was connected to a pump by rubber tubes. Different tube diameters and pump speeds were evaluated, and the assay was designed to mimic the half-lives of the antibiotics in vivo, i.e., 11.5 h for azithromycin, 5 h for clarithromycin, and 1 h for amoxicillin. Monolayers of HEp-2 cells were grown in the inserts for 2 days, after which H. pylori (clinical strain 88-23), was added to the system. Internalization was allowed for 12 h, and extracellular H. pylori cells were eradicated with gentamicin. The inserts were moved to the glass chamber, containing medium with 12.5 mg of either amoxicillin or azithromycin per liter or 2.4 mg of clarithromycin per liter. This represents 12.5, 50, and 80 times the extracellular minimum bactericidal concentration value, respectively. Samples were taken at 0, 2, 4, 6, 8, and 24 h. The HEp-2 cells were lysed, and intracellular bacteria were counted by plating. Inserts with infected cells grown in drug-free medium were included as controls for each time interval. A 3-log10 reduction of H. pylori was achieved in the experiments with azithromycin, and a 4-log10 reduction was achieved in the clarithromycin experiments, while no intracellular effect was seen when amoxicillin was used. The antibiotic concentrations at the sampling intervals were 12.5, 3.1, 0.8, 0.2, 0.05, and 0 mg/liter for amoxicillin; 12.5, 11.5, 10, 9, 8, and 3 mg/liter for azithromycin; and 2.4, 1.8, 1.4, 1, 0.8, and 0 mg/liter for clarithromycin. This new model for pharmacokinetic studies provides a useful tool, with applications for a broad range of microorganisms.

Amoxicillin↗
PubMed1998

Proarrhythmic and antiarrhythmic effects of bupivacaine in an in vitro model of myocardial ischemia and reperfusion.

BACKGROUND: Bupivacaine may have toxic cardiovascular effects when accidentally administered by intravascular injection. However, its electrophysiologic effects in the presence of myocardial ischemia remain unknown. The authors evaluated the electrophysiologic and anti- and proarrhythmic effects of bupivacaine in an in vitro model of the ischemic and reperfused myocardium. METHODS: In a double-chamber bath, a guinea pig right ventricular muscle strip was subjected partly to normal conditions and partly to simulated ischemia followed by reperfusion. The electrophysiologic effects of bupivacaine were studied at 1, 5, and 10 microM concentrations. RESULTS: Bupivacaine (5 and 10 microM) decreased the maximal upstroke velocity of the action potential (Vmax) in normoxic conditions and further decreased (10 microM) the Vmax decrease induced by ischemic conditions. Bupivacaine reduced the mean occurrence time to the onset of myocardial conduction blocks (9 +/- 3 min; mean +/- SD; P < 0.005 with 5 and 10 microM, compared with 17 +/- 6 min during simulated ischemia with no drug or control), and it increased the number of preparations that became inexcitable to pacing (55% of preparations, with 1 microM and 100% with 5 and 10 microM, compared with 17% for the control group). The incidence of spontaneous arrhythmias was reduced by 5 and 10 microM bupivacaine during ischemia and reperfusion and was enhanced by 1 microM bupivacaine during the ischemic phase. CONCLUSIONS: In guinea pig myocardium under ischemic conditions, bupivacaine induced a loss of excitability at concentrations of 5 and 10 microM. Proarrhythmic effects observed at 1 microM were considered as lower than the cardiotoxic range in normoxic conditions. The incidence of reperfusion arrhythmias was decreased at all concentrations.

Action Potentials↗
PubMed1998

Effect of propofol on oxidative stress in an in vitro model of anoxia-reoxygenation in the rat brain.

Propofol, an intravenous anaesthetic, is similar in chemical structure to the active nucleus of antioxidant substances such as alpha-tocopherol (vitamin E). The present study was designed to test whether propofol had antioxidant effects in an in vitro model of anoxia-reoxygenation in slices of rat brain. We used seven experimental groups: (1) control oxygenated tissue; (2) tissue subjected to anoxia for 20 min and reoxygenation for 3 h; and tissues processed as described and incubated with (3) Intralipid (commercial solvent for propofol), or propofol at a concentration of (4) 10 micromol/l, (5) 50 micromol/l, (6) 150 micromol/l or (7) 300 micromol/l. The production of lipid peroxides was quantified as thiobarbituric acid reactive substances (TBARS); tissular glutathione production and the activities of glutathione peroxidase (GSHpx), glutathione reductase (GSSGrd) and glutathione transferase (GSHtf) were also measured. Reoxygenation led to tissular oxidative stress, which was curtailed by propofol. The anaesthetic led to a 47% reduction in TBARS, a 165% increase in the reperfusion-inhibiting glutathione content, a 47% decrease in GSHpx activity, and an 87% increase in GSHtf activity. Intralipid had no effect on any of the parameters studied here. We conclude that propofol has a clear antioxidant effect in rat brain tissue subjected to anoxia-reoxygenation.

Animals↗
PubMed2004

Tools for anti-inflammatory drug design: in vitro models of leukocyte migration.

Inhibiting leukocyte recruitment is now a major focus in the design of novel anti-inflammatory drugs. Following the identification of lead compounds from conventional high-throughput screens using appropriate receptors or enzymes, it is important to validate the action of the compounds in a suitable in vitro model of leukocyte migration. Here, we review a range of different experimental approaches to modelling leukocyte migration, and identify the multi-well filter migration assay as the best compromise between the amount of resources required to screen multiple compounds and the amount of information gained about the effects of the compounds on cell movement behavior. However, there are pitfalls in the interpretation of data obtained using the multi-well filter migration assay, which arise from the imperfect correlation between the number of cells undergoing migration and the inhibitory activity of the test substances. We examine a number of such pitfalls and provide practical approaches to mitigate these problems as far as possible. We recommend a general strategy for screening inhibitors of cell migration using in vitro functional assays. While being more resource intensive than surrogate measures such as calcium flux, functional approaches nevertheless provide superior correlations with anti-inflammatory activity in vivo.

Anti-Inflammatory Agents↗
PubMed2004

Cytokines, nitric oxide, and cGMP modulate the permeability of an in vitro model of the human blood-brain barrier.

The endothelial cells (EC) of the microvasculature in the brain form the anatomical basis of the blood-brain barrier (BBB). In the present study, the effects of agents that modify the permeability of a well-established in vitro model of the human BBB were studied. The monolayers formed by confluent human brain microvessel endothelial cell (HBMEC) cultures are impermeable to the macromolecule tracer horseradish peroxidase (HRP) and have high electrical resistance. Exposure of HBMEC to various cytokines including TNF-alpha, IL-1beta, interferon gamma (IFN-gamma), or lipopolysaccharide (LPS) decreased transendothelial electrical resistance (TEER) mainly by increasing the permeability of the tight junctions. Primary cultures of HBMEC express endothelial nitric oxide synthase (eNOS) and produce low levels of NO. Treatment with the NO donors sodium nitroprusside (SNP) and DETA NONOate or the cGMP agonist 8-Br-cGMP significantly increased monolayer resistance. Conversely, inhibition of soluble guanylyl cyclase with ODQ rapidly decreased the resistance, and pretreatment of HBMEC with Rp-8-CPT-cGMPS, an inhibitor of cGMP-dependent protein kinase, partially prevented the 8-Br-cGMP-induced increase in resistance. Furthermore, NO donors and 8-Br-cGMP could also reverse the increased permeability of the monolayers induced by IL-1beta, IFN-gamma, and LPS. These results indicate that NO can decrease the permeability of the human BBB through a mechanism at least partly dependent on cGMP production and cGMP-dependent protein kinase activation.

Blood-Brain Barrier↗
PubMed2004

Factors inducing in-stent restenosis: an in-vitro model.

In-stent restenosis is caused by the proliferation of the smooth muscle cells (SMCs) following a host response towards the implanted device. However, the precise biochemical and cellular mechanisms are still not completely understood. In this paper, the behaviour of SMCs has been investigated by an in vitro model where the cells were stimulated by platelet derived growth factor (PDGF) on tissue-like substrates as well as on biomaterials such as stainless steel (St) and diamond-like carbon (DLC)-coated St. The results demonstrated that SMCs have a completely different adhesion mode on St and become particularly prone to proliferation and pro-inflammatory cytokine secretion under PDGF stimulus. This would suggest that restenosis may caused by the accidental contact of the SMC with the St substrate under an inflammatory insult.

Carbon↗
PubMed1990

Lack of a dopamine autoreceptor selective profile of B-HT 920 in functional in vitro model systems of D2 receptors in rat striatum.

Based on the results of in vivo studies, the thiazoloazepine derivative B-HT 920 has been proposed to be a selective agonist of dopamine autoreceptors. In the present study, we investigated the effects of B-HT 920 in two functional in vitro model systems of D2 receptors and compared these effects with the effects of the classical D2 agonist LY 171555. B-HT 920 and LY 171555 concentration dependently inhibited the electrically evoked release of radiolabeled dopamine and acetylcholine and the forskolin-induced stimulation of adenylate cyclase activity in rat striatal tissue slices with comparable efficacies. In striatal tissue slices prepared after 6-hydroxydopamine-induced destruction of dopaminergic terminals, both drugs were still able to inhibit forskolin-stimulated adenylate cyclase activity with a efficacy similar to that in tissue obtained from unlesioned rats. It is concluded that, in vitro, B-HT 920 is an agonist at both presynaptic and 'normosensitive' postsynaptic D2 receptors showing relatively high intrinsic activity.

Adenylyl Cyclases↗
PubMed1996

Interaction of LS174T human colon cancer cell mucins with Entamoeba histolytica: an in vitro model for colonic disease.

BACKGROUND & AIMS: Colonic mucins secreted by goblet cells protect the colon by preventing the attachment of enteric pathogens to the epithelium. Entamoeba histolytica overcomes this protective barrier and causes ulcerations, allowing the parasite to disseminate to the liver and form abscesses. An in vitro model is used to study the interaction between E. histolytica and colonic mucins. METHODS: Secretory mucins from the colonic adenocarcinoma cell line LS174T were collected and their functions assessed by their ability to inhibit amebic adherence to target cells and killing. The cytoprotective effect of mucus against E. histolytica cytolysis of LS174T monolayers was studied at 37 degrees C. RESULTS: Sepharose 4B column chromatography, metabolic labeling with [3H]glucosamine, cesium chloride density gradient centrifugation, and amino acid and carbohydrate compositional analysis revealed that LS174T cell mucins were typical of native colonic mucins. Mucin O-linked oligosaccharides bound to and inhibited the adherence of amebae to Chinese hamster ovary cells. E. histolytica killing of Chinese hamster ovary cell monolayers occurred rapidly, whereas killing of LS174T monolayers with an intact mucus layer was significantly retarded. CONCLUSIONS: Our results show that colonic mucins serve as the first line of host defense against amebic invasion and provide a useful model to study pathogen-mucin interactions.

Animals↗
PubMed1999

Effects of calcitonin on animal and in vitro models of skeletal metabolism.

During the 40 years since its discovery, calcitonin (CT) has been regarded primarily as an inhibitor of bone resorption and its therapeutic applications have been based on this property. A significant body of literature also indicates additional anabolic effects in animal and in vitro models. In a variety of bone loss histomorphometric models in the rat, CT, especially the salmon species, prevents or retards bone loss. In other species, similar results have been obtained, except in the beagle given human CT, in which a recent study reported increased bone resorption and bone loss. Consonant with the histomorphometric effects in several different species, bone mass (density) measured by a variety of methods increases, reversing the bone loss induced by the model. In related studies of mechanical properties, bone strength is increased by CT except in the beagle study which utilized human CT. In other species, experimentally induced fractures show either accelerated healing or heal normally, and there is no effect of CT to impair healing. Finally, studies of bone formation/mineralization strongly suggest an anabolic effect on cartilage formation, bone matrix synthetic activity, and bone growth. These animal effects are reflected by recent fracture prevention studies in humans. If its anabolic effects are ultimately found to be separable and additive to CT's basic action to inhibit bone resorption, new approaches to osteoporosis prevention, and possibly other treatment situations such as cartilage regeneration, may evolve using novel CT-like molecules.

Animals↗
PubMed2003

Development of an in vitro model to study oxidative DNA damage in human coronary artery endothelial cells.

The endothelial cell layer is a multifunctional barrier between the blood stream and the vascular wall. Reactive oxygen species can damage endothelial cells and may so potentiate atherosclerotic lesion formation. Therefore, we established an in vitro model for the qualitative and quantitative investigation of oxidative DNA damage and repair in human coronary arterial endothelial cells (CAEC). Oxidative DNA damage was induced by standardised treatment with 50-400 microM hydrogen peroxide (H(2)O(2)). The amount of DNA damage was quantified by determination of DNA single strand breaks (SSB) and alkali-labile sites in individual cells, using the alkaline single-cell gel electrophoresis assay--"comet assay". Significant DNA damage could be induced reproducibly in CAEC cells after exposure to 50 microM H(2)O(2) for 10 min. Additional treatment with catalase prevented DNA damage by H(2)O(2). The time needed for DNA repair depended on the initial grade of damage. After 30 min post-incubation at 37 degrees C, DNA damage was completely repaired in cells treated with 50 microM H(2)O(2). Cell samples initially damaged with H(2)O(2) concentrations between 100 microM and 400 microM were repaired after 60 min. This endothelial cell culture model allows experiments on oxidative DNA damage, alteration, and repair directly on the relevant target cells. Animals are used neither as direct objects of such experiments nor as cell or tissue donors.

Adult↗
PubMed1997

The effects of ion implantation on rate of tooth movement: an in vitro model.

Recently, the ion-implantation process has been applied to orthodontic wires. By altering the surface composition of a wire, the ion-implantation process supposedly decreases the frictional forces produced during tooth movement. The purpose of this study was to compare the amount of tooth movement produced by different orthodontic wire compositions, under identical conditions, by using an in vitro model. The wires tested were stainless steel, nickel-titanium (control and ion implanted), and beta-titanium (control and ion implanted). The amount of tooth movement was measured and compared. Results demonstrate that, stainless steel produced the least frictional force during in vitro tooth movement, followed by ion-implanted nickel-titanium, ion-implanted beta-titanium, untreated nickel-titanium, and finally, untreated beta-titanium. A Wilcoxon rank sum test showed statistically significant differences in the amount of movement seen with the ion-implanted wires when compared with their untreated counterparts.

Analysis of Variance↗
PubMed1989

[Alternative methods: development and use of two "in vitro" models for endocrine studies].

Most endocrinological studies are normally performed on several animal groups: one group for the determination of hormonal levels in tissues and plasma under basal conditions, a second group for the same determinations after "in vivo" treatment of the animals (stimulation or inhibition of the endocrine activities) and two additional groups for morphological investigations. Surgical approaches belong not rarely to the "in vivo" treatment of the animals. Moreover, the results from the different animal groups have to be extrapolated. In order to overcome these drawbacks, we have developed two "in vitro" models by means of which we can study on the tissues from the same animal: 1. function and structure of the hypothalamus and 2. function and structure of isolated pituitary cells. By using these models we can considerably reduce the number of animals needed for the studies, replace the "in vivo" by the "in vitro" experiments and refine the methods avoiding, among others, the extrapolation of results.

Animals↗
PubMed2006

Isolation and culture of murine heart and lung endothelial cells for in vitro model systems.

The inflammatory response is a critical component of host defense. An important goal of our group has been to understand the endothelial-dependent mechanisms that mediate leukocyte recruitment during an inflammatory response. In this chapter, we present a detailed method for the isolation and in vitro culture of murine vascular endothelial cells from the lung and heart. The endothelial cells are of high purity (85-99%) and retain certain of their functional differences, including constitutive and cytokine inducible adhesion molecule expression and chemokine production. Such endothelial cells, therefore, can be used for various in vitro models including leukocyte adhesion assay or in depth biochemical analyses.

Animals↗
PubMed1990

Pulmonary fibroblasts: an in vitro model of emphysema. Regulation of elastin gene expression.

Disruption and degradation of interstitial elastic fibers are significant characteristics of pulmonary emphysema. In order to examine the responses of elastogenic cells to the conditions mimicking degradation of interstitial pulmonary elastin, rat pulmonary fibroblast cultures were used as an in vitro model. Second passage fibroblasts were divided into two different environmental situations to represent cells adjacent to and remote from the site of elastase-digested matrix. One set of cell cultures was briefly digested with pancreatic elastase. The resultant digest was then added back incrementally to the medium of elastase-digested cell cultures and to the medium of a second set of undigested cultures. Both sets of cell cultures remained viable and metabolically active during these treatments (96 h of incubation) as judged by protein synthesis, cell number, and steady-state levels of beta-actin mRNA. However, the two sets of cultures exhibited opposite responses in elastin gene expression with addition of increasing amounts of the elastase digest. The elastase-digested cultures exhibited a 200% increase in extractable soluble elastin and a 186% increase in tropoelastin mRNA with the addition of increasing amounts of the elastase digest to the medium. Conversely, the amount of soluble elastin recovered from the undigested cultures decreased 75%, and the steady-state level of tropoelastin mRNA decreased 63%. Soluble elastin peptides generated from oxalic acid treatment of purified elastin were shown to decrease tropoelastin mRNA in undigested cell cultures in the same manner as the elastase digest. Based on these data, we propose that pulmonary fibroblast elastin gene expression can be controlled coordinately by the state of the extracellular matrix and solubilized peptides derived from that matrix. Such integrated regulation may serve to localize elastin repair mechanisms.

Actins↗
PubMed2000

In vitro model of the human liver parenchyma to study hepatotoxic side effects by Dy-EOB-DTPA.

RATIONALE AND OBJECTIVES: In vivo studies have shown species-specific toxicity after application of the liver-specific contrast agent Dy-ethoxybenzyl (EOB)-DTPA. To predict species differences in the laboratory, an in vitro model of the liver was used to examine the divergent results. METHODS: Rat, canine, porcine, and human hepatocytes were isolated and embedded between layers of collagen. During and after 48 hours of incubation with different concentrations of Dy-EOB-DTPA (maximum concentration 50 mmol/L), morphological changes and enzyme leakage were determined. RESULTS: The response to the contrast agent varied for hepatocytes from different species. For canine cells, morphological changes and cell death were evident with as little as 5 mmol/L Dy-EOB-DTPA. Rat hepatocytes tolerated up to 50 mmol/L Dy-EOB-DTPA, and enzyme leakage was transient. Only after incubation with 50 mmol/L Dy-EOB-DTPA was the formation of intracellular vacuoles evident. In contrast, even the highest concentration of Dy-EOB-DTPA did not cause an enzyme leakage of porcine or human hepatocytes, although similar vacuoles were seen. CONCLUSIONS: These data demonstrate a species-dependent toxicity for Dy-EOB-DTPA in vitro, with similar responses in porcine and human hepatocytes.

Animals↗
PubMed1994

An in vitro model for the effects of estrogen on neurons employing estrogen receptor-transfected PC12 cells.

Estrogen alters neurite outgrowth, neuritic spine development, and synaptogenesis in estrogen-responsive areas of the rat brain. However, examination of the specific effects of estrogen on neurons in vivo has been difficult. An in vitro model for the effects of estrogen on neurons was developed, using the PC12 rat pheochromocytoma cell line. Wild-type cells (PC12-WT) were stably transfected either with an expression vector coding for the full-length cDNA for the human estrogen receptor (hER), or with a control vector. Resultant clones were isolated, screened for incorporation of vector and expression of ER mRNA and protein, and analyzed for morphologic responses to estrogen. PC12-WT, NEO9 (ER-negative), and SER8 (ER-positive) cells exposed to 100 ng/ml NGF exhibited dose-responsive neurite outgrowth within 2 d by light microscopy (LM). Coadministration of 10(-10) to 10(-9) M estradiol (E2) had minimal effects on neurite outgrowth, neuritic spine development, or interneuritic connections in NEO9 or PC12-WT cells, but in SER8 cells E2 led to additive and dose-dependent increases in neurite outgrowth, spine development, and interneuritic connectivity. Coincubation of SER8 cells with E2 and the antiestrogen ICI 164,384 negated estrogenic effects on spine development and interneuritic connectivity. At the electron microscopic (EM) level, intercellular abutments of NEO9 or PC12-WT cells contained few and rudimentary gap junctions, with no increase by E2. However, SER8 cells exhibited augmented basal frequencies of gap junctions that increased with E2 incubation. Microinjection of Lucifer yellow into PC12-WT and NEO9 cells demonstrated low frequencies of dye coupling and no change with E2, but SER8 cells demonstrated increased dye-coupling frequency with E2 coincubation. The results suggest that SER8 cells recapitulate estrogen effects on neurons in vivo. Estrogen appears to induce an inherent neural morphologic program in estrogen receptor (ER)-containing cells. These three cell lines provide a unique in vitro system for studying mechanisms of estrogen-neuron interactions.

Animals↗