Circulating immunoreactivities of gastrin,glucagon and VIP after jejuno-ileal bypass

Seven Sprague-Dawley rats (404–440 g) underwent a 90% jejuno-ileal bypass (JIB); the functional loop consisted of $\text{1}\text{3}$ ileum and $\text{2}\text{3}$ jejunum with the bypassed loop being anastamosed to the ascending colon. Seven control rats were sham-operated. After 35 days, the rats were fasted 18 hours and venous blood was collected. Immunoreactivity of gastrin, measured with an antibody binding equally to G17 and G34, was higher in the plasma of the JIB (256±55 SEM pg/ml) than control (85±9 pg/ml) rats. This agrees with recent human studies but is in conflict with results in less mature rats. VIP levels were not significantly different. Glucagon-like immunoreactivity measured with antibodies specific for the C- and N-terminal regions of the hormone, respectively, were also higher in the JIB (510±40 and 129±15 pg/ml) rats.     

Denovo synthesis of prolactin by human decidua

Relatively large amounts of immunoreactive prolactin were measured in homogenates of human decidual tissue obtained immediately after delivery of normal term pregnancies. In order to study the release and possible synthesis of prolactin by this tissue, explants of decidua were incubated for 24 hours at 37°C in oxygenated Gey's buffer containing 20% fetal calf serum. When cycloheximide was added to the medium in concentrations sufficient to prevent $\text{in}$$\text{vitro}$ protein synthesis, 85–90% of the prolactin present in the tissue was released into the medium during the first 3 hours of incubation. No additional prolactin accumulated in either the medium or the tissue during the remainder of the incubation period. In the absence of cycloheximide, the prolactin concentration in the medium increased progressively during incubation, so that after 24 hours the total amount of hormone present in the tissue and medium was significantly greater than that in the tissue and medium prior to incubation (37.6 ± 9.6 ng/ml at 0 time vs 82.2 ± 7.7 ng/ml at 24 hours). When ³H-1-leucine (100 u Ci) was supplied during incubation, radioactive proteins were detected in the medium at 24 hr, 14–20% of which were specifically precipitated by antiserum to human pituitary prolactin. When aliquots of this medium were chromatographed on Sephadex G-100, 80–95% of the ³H-proteins precipitated by antiserum to pituitary prolactin eluted in the same position as did purified, iodinated pituitary prolactin. These data indicate that a species of prolactin which is identical to pituitary prolactin by the criteria of immunoprecipitation and gel chromatography is synthesized by human decidual tissue $\text{in}$$\text{vitro}$.     

The aging leydig cell: VII. Cytoplasmic estradiol receptors

Plasma estradiol and cytosolic estradiol receptor levels of testes were determined in a group of young (2–3 months) and old (24 months) Sprague-Dawley rats. Estradiol binding sites for the young rats averaged 5.6 ± 0.3 fmol/mg protein ($\text{x}\text{±}\text{SE, n}\text{=12}$), which was comparable to that of the old rats, 5.7 ± 0.3 fmol/mg protein (n=12). Using Scatchard analyses, the association constants at equilibrium of estradiol receptor binding of the old and young rats were the same, 6.1 × 10¹⁰M^?1. Plasma estradiol levels were also similar in both groups-19.6 ± 2.8 pg/ ml (n=14) for the young and 19.2 ± 2.6 pg/ml (n=10) for the old rats. Our results suggest that impaired testosterone biosynthesis in old rats was not due to elevated plasma estradiol levels or to differences in testicular estradiol receptor content.     

Elevation of plasma tyrosine after a single oral dose of L-tyrosine.

Plasma tyrosine concentrations in twelve normal, fasting human subjects were significantly elevated 2–8 hours after they ingested 100 mg/kg or 150 mg/kg tyrosine. Mean plasma tyrosine levels were maximal after 2 hours, rising from $\text{69 ± 3.9}\text{to}\text{154 ± 9.5}\text{nmols/ml}\text{(}\text{X}\text{±}\text{SEM}\text{)}$ after the 100 mg/kg dose and to 203 ± 31.5 nmols/ml after the 150 mg/kg dose (p ≤ 0.001 for both doses). The mean tyrosine ratio (defined as the ratio of plasma tyrosine concentration to the sum of the concentrations of six other neutral amino acids that compete for the same blood-brain barrier uptake system) increased from $\text{0.10 ± 0.02}\text{to}\text{0.28 ± 0.04 (}\text{X}\text{±}\text{SEM}\text{)}$ 2 hours after the 100 mg/kg dose (p ≤ 0.001) and to 0.35 ± 0.05 2 hours after the 150 mg/kg dose (p ≤ 0.005). No side effects of orally-administered L-tyrosine were noted.     

B-ETA-Endorphin immunoreactivity in rat and human blood: radioimmunoassay, comparative levels and physiological alterations.

Antiserum against human β-Endorphin (β_hEP) has been obtained from rabbit. The antiserum, diluted $\text{1}\text{1500}$ bound I¹²⁵ β_h-EP, demonstrating an effective range from 10pM to 10nM. The sensitivity of the assay is 2–3 fmoles. This antibody exhibits 10–15% cross-reactivity with human β-Lipotropin (β_h-LPH). β-EP-like immunoreactivity in rat blood has been detected in unextracted samples when compared to blood from hypophysectomized rats. The whole assay and calibration curves are carried out in plasma from hypophysectomized animals. β-EP-like immunoreactivity can be detected in normal rat plasma (75 ± 15 fmole/ml), and exhibits substantial increases with adrenalectomy (287 ± 32 fmoles/ml). In contrast, samples from five healthy normal human males gave values near the limits of detection of the assay (12 fmoles ± 3.9 per ml of plasma). Such values may be due to cross-reactivity of the antiserum with β_h-LPH or other circulating hormones. In contrast, patients with elevated ACTH production and normal pregnant humans exhibit significantly elevated levels of β-EP immunoreactivity in plasma.     

Enteric release of vasoactive intestinal peptide after a peptone meal in the dog

Vasoactive intestinal peptide (VIP) concentrations were measured by radioimmunoassay in plasma from portal and peripheral venous blood obtained from six alert, non-anesthetized dogs before and after gastric infusion of a 10% peptone meal. Mean basal portal and cephalic vein plasma VIP concentrations were $\text{42 ± 11.7}$ and $\text{42 ± 8.0}\text{(S.E.M.) pg/ml}$, respectively. No significant changes in peripheral venous plasma VIP concentrations were noted after the peptone meal throughout the duration of the collection period. In contrast, however, the mean VIP concentration in portal plasma increased promptly after the peptone meal with a peak of $\text{79 ± 8.2}\text{pg/ml}$ ($\text{P < 0.02}$) occurring 8 min after infusion of the meal. This was followed by a gradual decline in portal plasma VIP levels, with a return to prefeeding concentrations at 60 min ($\text{44 ± 6.3}\text{pg/ml}$). Results of these studies demonstrate that following gastric infusion of a peptone meal in the dog, portal, but not peripheral, plasma VIP concentrations increase significantly. Failure to detect augmentation of peripheral vein VIP levels after the meal is probably due to hepatic clearance of VIP.     

Measurement of thromboxane B2 in human plasma or serum by radioimmunoassay

A radioimmunoassay for thromboxane B₂ (TxB₂), a stable metabolite of thromboxane A₂, is described. The method consists of extraction of TxB₂ into ethyl acetate from acidified plasma or serum samples and saturation analysis using specific antibodies produced in rabbits against TxB₂-BSA conjugate. The 50 % displacement level of the standard curve was 19.1 ± 2.9 pg/tube (mean ± S.D., n = 19). The method blank was 3.4 ± 3.1 pg/ml (n = 15) and the assay sensitivity thus 9.6 pg/ml (mean blank + 2 S.D.). When 100 to 200 pg of TxB₂ were added to plasma, 96.2–103.6 % were recovered. The intra-assay coefficient of variation varied from 6.7 to 9.7 %, and the inter-assay coefficient of variation was 18.6 % (n = 10). The TxB₂ concentration in the plasma of 14 healthy subjects varied from 29.3 to 120.8 pg/ml with a mean ± S.D. of 70.1 ± 26.1 pg/ml, when the blood was collected into tubes containing acetylsalicylic acid (ASA), whereas significantly higher (p < 0.001) TxB₂ concentrations of 68.3 – 285.3 pg/ml with a mean ± S.D. of 151.8 ± 66.6 pg/ml were obtained from the same subjects in the plasma of blood which was collected into tubes containing no ASA. When blood samples from 10 subjects were allowed to clot at 0, +24 or +37°C for 60 min., the TxB₂ concentrations in the sera were 2053 ± 870 pg/ml, 4001 ± 1370 pg/ml and 178557 ± 54000 pg/ml, respectively. The TxB₂ levels in sera which were separated from blood samples incubated at +37°C, correlated significantly (p < 0.001) with the TxB₂ productions in platelet-rich-plasma (PRP) after an induced aggregation. Our results indicate 1) when TxB₂ is measured in plasma, the use of prostaglandin synthesis inhibitor in the collection tubes is necessary and 2) the measurement of TxB₂ in serum of blood which has been kept at +37°C for a strictly standardized period of time could replace the use of PRP in TxB₂ studies.     

Cellular pH and water permeability control in frog urinary bladder a possible action on the water pathway

Mucosal acidification (from pH 8.1 to 6.0) reversibly inhibited the hydroosmotic responses to oxytocin, cyclic AMP and 8-bromo-cyclic AMP in frog urinary bladder. These inhibitory effects were only observed in the presence of a permeant buffer in the apical medium and could also be elicited by CO₂ bubbling, even when the mucosal pH was clamped at 8.1. Acid pH reduced the oxytocin-induced net water flux faster than norepinephrine or oxytocin removal and the difference was especially important at low temperature. The time course of recovery from acid pH inhibition was, at 20°C, similar to that of the hormonal action, but when the medium temperature was reduced to 6–7°C, the recovery from acid pH inhibition paradoxically became faster while the oxytocin action was markedly slowed down ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of changes in net water fluxes (expressed in min): oxytocin addition at 20°C, $\text{6.2 ± 0.9}$; at 6°C, $\text{24 ± 3}$; oxytocin removal at 20°C, $\text{4.7 ± 0.8}$; at 6°C, $\text{22 ± 3}$; pH inhibition at 20°C, $\text{2.6 ± 0.2}$; at 6°C $\text{2.5 ± 0.2}$; recovery from pH 6 at 20°C, $\text{6.5 ± 0.9}$; at 6°C, $\text{2.7 ± 0.3}$). These results can be explained by accepting two main loci sensitive to medium acidification: (1) the cyclase system and (2) an intracellular, temperature-independent, post-cyclic AMP site. The fact that the intramembranous particle aggregates associated with the oxytocin-induced water permeability increase did not disappear after the flow inhibition by acid pH at low temperature suggests that the second effect could be located at the water channel itself.     

18,19-Dihydroxydeoxycorticosterone; A novel product of cytochrome P-45011β-catalyzed reaction

After incubating 18-hydroxydeoxycorticosterone (18-OH-DOC) with cytochrome P-450_11β in the reconstituted system, the products were analyzed with HPLC. There appeared two product-peaks on the chromatogram, one of which was identified as a peak of 18-hydroxycorticosterone (18-OH-B), an expected product of the 11β-hydroxylation. Another peak did not coincide with those of any known corticoids. This unidentified product was further purified, and the purified material was analyzed by gas chromatography-mass spectrometry ($\text{GC}\text{MS}$). The mass spectrum showed that the unidentified product is one of the structural isomers of 18-OH-B. A further analysis with ¹H-NMR spectrometry indicated that a proton resonance peak of 19-CH₃ in 18-OH-DOC disappeared in the product and the methyl group of the substrate seemed to be converted to -CH₂OH. These results suggested that the unidentified product generated from 18-OH-DOC by P-450_11β-linked hydroxylase system may be 18,19-dihydroxydeoxycorticosterone (18,19,21-trihydroxypregn-4-ene-3,20-dione; 18,19-diOH-DOC), a hitherto unreported corticoid.     

Renal metabolism of substance P in the dog.

Substance P is a potent vasodilatory, diuretic, and natriuretic agent. Since subcellular fractions of the kidney rapidly inactivate substance P in vitro, the present study was designed to examine this observation $\text{in}$$\text{vivo}$ in anesthetized dogs. Arterial, renal venous, and urinary levels of immunoreactive substance P were determined by radioimmunoassay and were found to be 117±11, 128±12 and 659±104 pg/ml, respectively. The urinary and fractional excretion of immunoreactive substance P were 122±22 pg/min and 6.6±2.0%, respectively. When substance P was infused intravenously, the arterial and renal venous plasma levels of immunoreactive material increased whereas the urinary levels did not change. Infusions of 50 ng/kg/min of substance P significantly decreased mean arterial pressure, urinary volume, creatinine clearance as well as the urinary excretion, clearance, and fractional excretion of immunoreactive substance P. During intrarenal infusion of ¹²⁵I-(8-Tyr) substance P, high levels of radioactive material were found in the urine and renal venous plasma which failed to migrate on thin layer chromatography with intact ¹²⁵I-(8-Tyr) substance P. Thus under these conditions, intact substance P was not released from the kidney into the urine or renal venous blood, but instead circulating substance P was rapidly and completely metabolized, probably by both vascular and tubular elements of the kidney.     

Solid phase radioimmunoassay of plasma aldosterone

A solid phase radioimmunpassay for the measurement of aldosterone in plasma is described. The antiserum was produced by immunizing rabbits with 3-carboxymethyloxime of aldosterone-18-21-diacetate coupled to bovine serum albumin, This antiserum was covalently linked to an iminocellulose according to the procedure of Wide and used in the assay at a $\text{1}\text{1050}$ final dilution. It contained antibodies with association-constant of 1.1 × 10¹⁰M^?1 and was fairly specific since with the exception of aldosterone acetates, none of the tested steroids cross-reacted more than 0.05 per cent.Aldosterone was extracted with dichloromethane, purified by paper chromatography, then submitted to the assay. The intra-assay reproducibility varied between 4 and 13 % and the inter-assay reproducibility between 13 and 21 %. The least detectable amount was 5 pg per tube. This method is very simple and, with the exception of the Chromatographie step, can be completed in half a working day.     

Solid phase radioimmunoassay of plasma aldosterone

A solid phase radioimmunoassay for the measurement of aldosterone in plasma is described. The antiserum was produced by immunizing rabbits with 3-carboxymethyloxime of aldosterone-18–21-diacetate coupled to bovine serum albumin. This antiserum was covalently linked to an iminocellulose according to the procedure of Wide and used in the assay at a $\text{1}\text{1050}$ final dilution. It contained antibodies with association-constant of 1.1 × 10¹⁰ M^?1 and was fairly specific since with the exception of aldosterone acetates, none of the tested steroids cross-reacted more than 0.05 per cent.Aldosterone was extracted with dichloromethane, purified by paper chromatography, then submitted to the assay. The intra-assay reproducibility varied between 4 and 13 % and the inter-assay reproducibility between 13 and 21 %. The least detectable amount was 5 pg per tube. This method is very simple and, with the exception of the chromatographic step, can be completed in half a working day.     

The falling drop method for deuterium oxide: Factors influencing accuracy

Factors affecting the accuracy of the falling drop method for D₂O were considered: selection of temperature of the constant-temperature bath, permissible temperature fluctuations, and D₂O concentration. Bath temperatures used, 27.24 to 27.32°, and constancy within ±0.002° had no influence on the regression relating drop velocity to concentration, v = a + bc, or sampling error (S.D._v). The latter increases significantly with concentration from 0.05 to 0.23 ml % D₂O, is significantly smaller than experimental error (S.D._e), and is inappropriate for estimating intra-assay limits. More appropriate is the S.D._c derived from the variance of the terms of the regression equation. Assay results using this error term can be expected to vary from about ±4.8% at 0.05 ml % to about ±2% at 0.23 ml %. These results compare favorably with those reported for the mass spectrograph used to determine the mass ratio of $\text{HDO}\text{H}{}_2\text{O}$ vapor. A more conservative estimate is obtained by using $\text{λ}\text{=}\text{S.D.}{}_{\mathrm{e}}\text{b}$, which in this work indicated within-assay variability of ±12.4% at 0.05 ml % and ±2.7% at 0.23 ml % D₂O. The term S.D._e, corresponding to 0.0062 ml % in these experiments, provides the best means for assay comparisons.Satisfactory recoveries of D₂O added to urine averaging 99% of known values were obtained after shell-freezing, without distillation of standards and without permanganate oxidation of samples. No increase in error beyond that anticipated from standards alone was observed when urine was the vehicle.     

Catecholamine-induced changes in transmembrane potential and temperature in normal and dystrophic hamster brown adipose tissue

Previous studies have shown that norepinephrine (NE) elicits trans-membrane potential changes in skeletal muscle cells from normal and dystrophic (BIO 14.6) hamsters, with the magnitude of these changes being significantly less in dystrophic cells. To determine if the decreased response of the dystrophic muscle cells reflects a more generalized phenomenon, the present study was designed to evaluate the effects of NE on membrane properties of brown adipocytes. $\text{In vivo}$ techniques using glass microelectrodes were similar to those used in the muscle studies. NE injection (2 to 5 μg/kg body wt, i.v.) into anesthetized hamsters was followed by membrane depolarization, the magnitude of which did not significantly differ in the dystrophic and normal adipocytes. For example, upon administration of 5 μg NE/kg body wt, the average depolarization was $\text{14.5 ± 1.3}\text{mV}\text{(}\text{X}\text{±}\text{S.E.}\text{)}$ for 20 dystrophic cells and 14.1 ± 1.8 mV for 18 normal cells. The depolarizations following i.v. infusion of isoproterenol and phenylephrine also had similar amplitudes in both normal and dystrophic cells. Despite this lack of difference in plasma membrane responses, NE induced a significantly smaller rise in interscapular brown fat temperature in the dystrophic (0.09°C) than in the normal hamsters (0.26°C) following administration of 5 μg NE/kg body wt. Thus, the decreased responsiveness to NE of dystrophic sarcolemma did not occur with the plasma membrane of brown adipocytes, although brown fat temperature changes in the dystrophic hamsters were decreased in amplitude.     

Induction kinetics of human suppressor cells in the mixed lymphocyte reaction and influence of prednisolone on their genesis

The induction kinetics of human suppressor cells in mixed lymphocyte cultures (MLC) and the influence of prednisolone on the genesis of these suppressor cells is reported. We induced over 1 to 6 days suppressor cells in one-way MLC (MLC-1), the inhibitory activity of which was tested on a secondary MLC (MLC-2), and on responder cells alone, where lymphocytes were obtained from the same lymphocyte donors as for the MLC-1. In four experiments the degree of inhibition ($\text{x}\text{?}\text{±}\text{SE}$) when suppressor cells were induced for 2, 4, or 6 days was 38.5 ± 11.8, 79.5 ± 7, and 85 ± 6%, respectively, compared to 50.5 ± 9.4, 83.3 ± 7.8, and 85.3 ± 9.8% when 500 ng/ml prednisolone was added to the MLC-1. A similar inhibition pattern was observed when the generated suppressor cells were incubated with responder cells only. The inhibitory activity of these MLC-induced suppressor cells was abrogated by irradiation with 3000 R. Suppressor cells apparently are generated in MLCs between Days 1 and 4; furthermore, their genesis is not affected by usual therapeutic concentrations of prednisolone.     

Effects of lighting programs on onset of the ovulatory season in mares

Using 240 pony mares, lighting regimens were tested for their efficiency in hastening the onset of the ovulatory season. The mean number of days from January 1 to first ovulation was used as the end point. No advantage was gained by beginning a fixed lighting regimen ($\text{15.5L}\text{8.5D}$, hours light/hours dark) November 1 (66 ±8) versus December 1 (65 ±9), but beginning on January 1 was less efficient (98 ±8; controls, 132 ±5; P<0.05). In another experiment, daily three-hour interruptions of either the light phase (67 ±10) or the dark phase (71 ±11) did not significantly retard the effectiveness of a fixed regimen of $\text{15L}\text{9D}$ (54 ±5; controls, 142 ±6). A $\text{15L}\text{9D}$ regimen every other day (natural day length on alternate days) resulted in an interval (85 ±7) that was shorter (P<0.05) than for the controls and longer (not significant) than for the daily $\text{15L}\text{9D}$ regimen. When used with natural day length, a one-hour pulse of light in the evening (15 hours after sunrise) was not effective (141 ±6); a one-hour pulse in the morning 9.5 hours after sunset) was only partially effective (117 ±6). In another experiment, the interval was reduced (P<0.05) in a group with one hour of light fixed at 4:00 a.m. with natural day length (85 ±8; $\text{15L}\text{9D}$, 75 ±7; controls, 126 ±9). Results indicated that a fixed one-hour pulse of light at 4 a.m., used with natural day length, may provide an acceptable level of stimulation.     

The effects of prostacyclin (PGI2) and 6-keto-PGF1α on bovine plasma progesterone and LH concentrations

Injections of 1 mg PGI₂ directly into the bovine corpus luteum significantly increased peripheral plasma progesterone concentrations within 5 min. Concentrations were higher in the PGI₂-treated heifers than in saline-injected controls between 5 and 150 min and at 3.5, 4, 5, and 7 h post-treatment. Levels tended to remain elevated through 14 h. Saline and 6-keto-PGF_1α were without effect on plasma progesterone levels. The luteotrophic effect of PGI₂ was not due to alterations in circulating LH concentrations. An $\text{in vitro}$ experiment assessed the effects of either PGI₂ alone or in combination with LH on progesterone production by dispersed luteal cells. Progesterone accumulation over 2 h for control, 5 ng LH, 1 μg PGI₂, 10 μg PGI₂, and 10 μg PGI₂ plus 5 ng LH averaged 99 ± 42, 353 ± 70, 152 ± 35, 252 ± 45, and 287 ± 66 ng/ml (n=4), respectively. Thus PGI₂ has luteotrophic effects on the bovine CL both $\text{in vivo}$ and $\text{in vitro}$.     

Liquid scintillation counting of aqueous samples using Triton-containing scintillants

Markedly unstable count rates were observed using a toluene-Triton (2:1, $\text{v}\text{v}$) scintillant during counting of water-soluble radioactive compounds when < 5% ($\text{v}\text{v}$) water was present, because of the separation of phases. Efficiency correction in these instances could not be made by using ³H₂O as internal standard, because under the same conditions count rates with tritiated water were stable. Increasing water to ≥6% stabilized the count rates. With toluene-Triton (2:1, $\text{v}\text{v}$) scintillant, the water level should preferably be maintained between either 6 and 12 or 18 and 24% for ¹⁴C- and ³H-labeled compounds for counting at 6°C or at ambient temperature (but only between 6 and 12% for ³H counting at room temperature). With a “Tritosol” (Anal. Biochem.63, 555 (1975) modified to contain 35 ml of ethylene glycol, 140 ml of ethanol, 250 ml of Triton X-100, 575 ml of xylene, 3 g of PPO, and ±200 mg of POPOP, water levels of up to 23% were acceptable for ¹⁴C and ³H for counting at room temperature or at 6°C. Within these limitations, with the toluene-Triton or with the modified Tritosol as scintillant, both polar and apolar radioactive compounds exhibited similar efficiencies and gave quench-correction curves, based on the external standard ratio, that were linear for both ¹⁴C and ³H-labeled compounds.     

Oral ingestion of insulin liposomes: Effects of the administration route

We prepared an insulin liposome suspension by hot dispersion (50 °C) of a lipid mixture comprising dipalmitoyl phosphatidylcholine (DPPC) and cholesterol (7:2 molar ratio) in an 80 UI/ml acid bovine insulin solution, followed by two minutes of cold sonification (4 °C). Free insulin was removed by ultracentrifugation and the washed insulin liposomes were resuspended in a 1% aqueous saline solution (pH 3). Administration of these liposomes in the buccal cavity of normal rats caused clear hypoglycemia (?37% of the initial glycemia after one hour and ?44% after $\text{2}\text{1}\text{2}$ hours), but the solution was inactive when introduced by a strictly intragastric route. Hypoglycemic effects were also obtained when a mixture containing a liposome suspension devoid of insulin and 10 UI/100 g b.w. of free insulin was given by the buccal route (?56% of initial glycemia one hour later and ?55% after $\text{2}\text{1}\text{2}$ hours). These results show that the route of liposomal insulin administration strongly influences its biological effects.     

Role of the plasma membrane in the mechanism of cholesterol efflux from cells

In order to investigate the role of the plasma membrane in determining the kinetics of removal of cholesterol from cells, the efflux of [³H]cholesterol from intact cells and plasma membrane vesicles has been compared. The release of cholesterol from cultures of Fu₅AH rat hepatoma and WIRL-3C rat liver cells to complexes of egg phosphatidylcholine (1 mg / ml) and human high-density apolipoprotein is first order with respect to concentration of cholesterol in the cells, with half-times ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$) for at least one-third of the cell cholesterol of 3.2 ± 0.6 and 14.3 ± 1.5 h, respectively. Plasma membrane vesicles (0.5–5.0 μm diameter) were produced from both cell lines by incubating the cells with 50 mM formaldehyde and 2 mM dithiothreitol for 90 min. The efflux of cholesterol from the isolated vesicles follows the same kinetics as the intact, parent cells: the $\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ values for plasma membrane vesicles of Fu₅AH and WIRL cells are 3.9 ± 0.5 and 11.2 ± 0.7 h, respectively. These $\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ values reflect the rate-limiting step in the cholesterol efflux process, which is the desorption of cholesterol molecules from the plasma membrane into the extracellular aqueous phase. The fact that intact cells and isolated plasma membranes release cholesterol at the same rate indicates that variations in the plasma membrane structure account for differences in the kinetics of cholesterol release from different cell types. In order to investigate the role of plasma membrane lipids, the kinetics of cholesterol desorption from small unilamellar vesicles prepared from the total lipid isolated from plasma membrane vesicles of Fu₅AH and WIRL cells were measured. Half-times of cholesterol release from plasma membrane lipid vesicles of Fu₅AH and WIRL cells were the same, with values of 3.1 ± 0.1 and 2.9 ± 0.2 h, respectively. Since bilayers formed from isolated plasma membrane lipids do not reproduce the kinetics of cholesterol efflux observed with the intact plasma membranes, it is likely that the local domain structure, as influenced by membrane proteins, is responsible for the differences in $\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ values for cholesterol efflux from these cell lines.