Atrial natriuretic peptide receptors on the corneal endothelium.

PURPOSE: To ascertain whether the corneal endothelium exhibits specific receptors for atrial natriuretic peptide (ANP) and to characterize partially any binding activity. METHODS: Most experiments employed radioligand-binding techniques to characterize 125I-ANP binding activity. Guanylate cyclase and corneal deturgescence assays were used in an attempt to correlate 125I-ANP binding activity with physiologic processes. RESULTS: Corneal endothelial cells reversibly bound 125I-ANP with high affinity and exhibited a finite number of 125I-ANP binding sites. These binding sites also bound several ANP fragments but did not show binding affinity for the peptide hormone, vasopressin. Autoradiograms of affinity labeled ANP receptors separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed specifically radiolabeled proteins of several molecular weights in nonreduced samples, but only one major radiolabeled protein of 65 kd when samples were chemically reduced before separation. CONCLUSIONS: Corneal endothelial cells from several species exhibit specific binding of 125I-ANP. The binding characteristics of these receptors are similar to physiologic ANP receptors identified in other tissues. Several lines of evidence indicate that corneal endothelial ANP receptors are the "clearance" (ANP-C) type.

Muscarinic cholinoceptors in native and cultured human corneal endothelium.

Specific and high affinity binding of the potent muscarinic cholinergic antagonist, [3H]quinuclidinylbenzilate ([3H]QNB) was observed using intact native and cultured adult human corneal endothelium (HCE). Specific binding was proportional to radioligand concentration between 0.03 and 5 nM, indicating a maximal binding capacity (Bmax) of 130 fmol of [3H]QNB/mg protein and a dissociation constant (Kd) of 0.3 nM. Atropine competed effectively with [3H]QNB for binding sites; requiring 3 nM to inhibit 50% of the binding of 1 nM [3H]QNB. Carbachol also competed with [3H]QNB at higher concentrations, but nicotine did not affect [3H]QNB binding at levels up to 1 nM. [3H]QNB binding was also observed in cultured cells of adult human, rabbit, and bovine corneal endothelium. Native and cultured HCE were affinity labelled using tritium-labelled propylbenzilylcholine mustard (PBCM). Separation of the proteins in affinity labelled native and cultured tissue by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) showed that only one protein in each preparation, of 60 and 55 kilodaltons (kDa), respectively, was specifically radiolabelled. These data indicate that the corneal endothelium of human and several animal species exhibit muscarinic cholinoceptors.

Alpha 1-adrenoceptors in the corneal endothelium.

The potent alpha 1-adrenoceptor antagonist, [3H]prazosin, exhibited high affinity, specific and reversible binding to intact rabbit, bovine and human corneal endothelial cells in culture. The binding of 1 nM [3H]prazosin to rabbit cells reached a steady-state level within 10 min at 37 degrees C. Under these conditions, approximately 50% of the [3H]prazosin bound was specific. The level of specific [3H]prazosin binding was concentration-dependent, but Rosenthal analysis indicated that [3H]prazosin bound to at least two sites. One site exhibited a high affinity for [3H]prazosin (Kd = 0.2 nM), but a relatively low binding capacity (Bmax = 175 fmol bound mg-1 protein); the other site showed a relatively low affinity for the radioligand (Kd = 85 nM), but a much higher binding capacity (1280 fmol mg-1). Several known alpha 1-adrenoceptor antagonists and agonists competitively inhibited [3H]prazosin binding at the high affinity site when incubated with the radioligand. The relative potencies of these competing ligands were generally consistent with their binding affinities for alpha 1-adrenoceptors in other tissues. Phenylephrine stimulated the rate of hydrolysis of phosphatidylinositol 4,5-bisphosphate to inositol 1,4,5-trisphosphate by 63% in these cells. This stimulation was inhibited by 52% if phentolamine was also present during the incubation. These data indicate that corneal endothelial cells have alpha 1-adrenoceptors which can modulate polyphosphoinositide turnover in this tissue.

The menarcheal age of Chinese girls.

The menarcheal age of Chinese girls was analysed based on a national survey in 1985. A total of 162,902 girls, aged 7-22 years, of Han nationality and 34,232 girls, aged 7-18, of 27 minority nationalities were sampled. Data were further subdivided into urban and rural areas. It was found that the curves of menstruating percentage in each of the subgroup were in long sigmoid shapes. The menstruating rate (MR) of Han girls was higher than that of the minority girls in almost every age group. The median menarcheal age (MMA) of Han girls was significantly lower than that of the minority girls. In Han nationality the urban girls showed higher MR and lower MMA than the rural girls. The influence of hereditary, cultural and economical factors on menarcheal age in Chinese girls is discussed. The data of this study were also compared with research reports from other world literature.

Corneal function after storage in dexsol or optisol.

Rabbit corneas were stored in Dexsol or Optisol (Chiron, Irvine, CA) for up to 2 wk at 4 degrees C. The thickness of corneas placed in Dexsol decreased 10 microns after they were placed initially in Dexsol, then increased approximately 8 microns/d for 7 d and 3 microns/d thereafter. Corneas placed in Optisol decreased 35 microns in thickness initially, then increased 2 microns/d thereafter. Human corneas showed similar thickness changes to those of the rabbit when stored in these media. After 5.5, 10, and 14 d in storage, rabbit corneas from each medium were cultured to assess their net deturgescence ability. Identical groups were cultured in media containing 20 microM ouabain to monitor the corneas' passive swelling characteristics. Corneas stored in either medium showed similar net deturgescence and passive swelling patterns after each storage period. Deturgescence rates decreased with increasing storage time, primarily because the rates of passive corneal swelling increased with storage time. Knowledge of the net deturgescence and passive swelling rates allowed an estimation of the total deturgescence activity of corneas after removal from Dexsol or Optisol. The total deturgescence activity of corneas stored in Dexsol for 5.5, 10, and 14 d was 85%, 68%, and 63% of control corneas, which were processed identically but not stored before culture. Corneas stored in Optisol exhibited 87%, 71%, and 69% of control deturgescence activity, respectively. These experiments show that Optisol was not significantly better than Dexsol in retaining poststorage corneal deturgescence activity but was superior to Dexsol in preventing corneal swelling during storage.

Secular change in the growth and development of Han children in China.

This paper reports the secular change in physical growth and development of Han children in 12 Chinese metropolitan cities including Beijing, Tianjing, Shanghai, Jinan, Harbin, Shenyang, Changchun, Nanjing, Hangzhou, Wuhan, Guangzhou, and Chengdu. Based on a recent national survey of 14,688 healthy schoolchildren, aged 7-18 years, and together using the historical records of healthy schoolchildren as secondary data source, this study found a secular change in the growth and development of Chinese children and youth. In every decade between the 1950s and 1985 the average height increased by 2.66 cm (range 1.78-3.77) for boys and 2.40 cm (range 1.72-3.76) for girls, and the average weight increased by 1.64 kg (range 1.28-2.63) for boys and 1.14 kg (range 0.63-2.01) for girls in the 12 cities. In the last decade from the 1970s to 1985, the height and weight increments were highest in Beijing, Jinan, Shenyang and Shanghai. In addition, the increments were higher during the peak growth years than during other periods of growth. In the 30 years studied, the ages of maximum growth velocity were advanced, and they were 1 or 2 years earlier in 1985 than in the 1950s in most cities. When compared with the data from the 1930s the results indicate that, during the last half-century, height increased by 1.12-2.66 cm per decade for boys and 1.42-2.67 cm/decade for girls, while weight increased by 0.56-1.27 for boys and 0.65-1.18 kg for girls in Beijing, Shanghai, Nanjing, Hanzhou and Guangzhou.

Physical growth of Chinese school children 7-18 years, in 1985.

Physical development in Chinese school children was analysed based on a national survey in 1985. Close to half a million children and adolescents, aged 7-18, from Han ethnic group and 27 minority ethnic groups were assessed for height, weight, sitting-height, chest circumference, biacromial and biiliac diameters. Growth in Han children was greater than that in minority ethnic group children. The growth differences among minority ethnic groups were large. Growth in urban children was greater than that in rural children except in a few age groups of girls. A significant difference in children's growth was found between northern and southern divisions of China, it being greater in the north. The growth curves of physical characteristics demonstrated systematic regularity. Finally, the results of this study were compared with research reports from other world literature.

The effects of UW solution and its components on corneal thickness during and after storage.

The ability of rabbit corneas to undergo energy-dependent deturgescence was examined after the corneas were stored at 4 degrees C in UW solution, M-K media, or selected modifications of these media. All corneas slowly increased in thickness during storage, despite the presence of colloidal osmotic agents. Corneas stored for 2.5 days in M-K became slightly thinner when cultured over a 24-hour period. Corneas stored in UW swelled quickly in culture and became too opaque to measure within three hours. Corneas stored in UW with 1.8 mM CaCl2 swelled transiently, then maintained their thickness and exhibited deturgescence in the latter stages of the culture period. Deturgescence of corneas stored for 7 days in M-K was only slightly worse than those stored for 2.5 days. Corneas stored for 7 days in UW, however, became opaque almost immediately in culture and those stored in calcium-supplemented UW became opaque within 4.5 hours. Replacement of the dextran in M-K with hydroxyethyl starch produced a slower rate of corneal swelling during storage and a substantially better corneal deturgescence profile during culture. Use of high concentrations of potassium ion in the M-K formulation had no significant effect on post-storage deturgescence. Replacement of glucose in M-K with the impermeable sugar, raffinose had a slight deleterious effect on corneal deturgescence in subsequent culture. Use of the impermeable anions gluconate or lactobionate to replace chloride ion caused profound corneal swelling during culture, compared with those stored in M-K. These experiments show that UW solution is inferior to M-K at preserving post-storage corneal function.(ABSTRACT TRUNCATED AT 250 WORDS)

Alpha 1-adrenoceptors in human corneal epithelium.

Specific binding of the potent, selective alpha 1-adrenoceptor antagonist 3H-prazosin was demonstrated in cultured human corneal epithelial cells. Specific binding of the radioligand was concentration-dependent between 0.5 and 6 nM, with apparent saturation of receptor sites seen at higher concentrations. The cells exhibited a maximum binding capacity for 3H-prazosin of 225 fmol/mg of cellular protein and a dissociation constant of 2 nM. The binding of 3H-prazosin was competitive with known alpha 1-adrenoceptor ligands and was reversible. Epithelium of intact human corneas also exhibited specific 3H-prazosin binding, as did cultures of bovine and rabbit corneal epithelium. The alpha-adrenergic agonist methoxamine significantly stimulated phosphatidylinositol 4,5-bisphosphate hydrolysis, measured as myoinositol trisphosphate accumulation in cultures of human corneal epithelium. This stimulation was inhibited by the presence of prazosin during the assays. These findings indicate the existence of specific, reversible, high-affinity receptors for alpha 1-adrenoceptors that regulate inositol phosphate turnover in human, rabbit, and bovine corneal epithelial cells.

Corneal function after storage in commercial eye bank media.

Rabbit corneas were stored in commercially prepared media (K-Sol, M-K, CSM, or Dexsol) for 3, 6, 9, or 12 days. Corneas stored in each medium showed decreased post-storage deturgescence with increased storage time. This effect was most pronounced in corneas stored in K-Sol, followed by those in M-K, CSM, and Dexsol. Corneas stored in K-Sol also exhibited the fastest swelling rate when cultured in the presence of 20 microM ouabain, followed by those in M-K, Dexsol, and CSM. An estimate of the active ion transport capacity of each experimental group was made by determining the area between each group's corneal thickness profiles in the presence and absence of ouabain. Corneas stored in M-K, K-Sol, or CSM retained approximately 70% of control activity after 3 days storage and 65% after 6 days storage. Corneas stored in Dexsol had 92% of control activity after 3 days of storage, 78% after 6 days, and 44% after 12 days.

Muscarinic cholinoceptor regulation of cyclic guanosine monophosphate in human corneal epithelium.

Cultured human corneal epithelial cells showed high-affinity, specific binding to the muscarinic cholinergic antagonist, 3H-quinuclidinyl benzilate (3H-QNB). The binding sites had a dissociation constant of 3.9 nM and a maximal binding capacity of 880 fmol bound/mg protein. Other muscarinic antagonists (cyclopentolate and atropine) effectively competed for binding with 3H-QNB at low concentrations (IC50 = 10 nM). The muscarinic cholinergic agonist carbachol also competed for binding with 3H-QNB at somewhat higher concentrations (IC50 = 0.5 microM), and the nicotinic cholinergic agonist, nicotine, was at least 400-fold less potent than carbachol. Carbachol stimulated cyclic guanosine monophosphate (GMP) levels in these cells up to threefold over control. This stimulation was sensitive to atropine inhibition, requiring only 2 nM atropine for 50% inhibition and 100 nM of atropine to block the carbachol effect completely. A 90% decrease in specific 3H-QNB binding was observed if cultured cells were homogenized and fractionated before assay. Significant levels of specific 3H-QNB binding could also be observed when intact human corneas were incubated with 3H-QNB and their epithelium subsequently isolated before measurement of bound radioligand. These studies indicate the presence of muscarinic cholinoceptors in human corneal epithelium which are associated with control of cyclic GMP levels in this tissue.

Muscarinic receptors and their regulation of cyclic GMP in corneal endothelial cells.

Particulate fractions from fresh bovine corneal endothelium exhibited high affinity, specific binding by a potent muscarinic cholinergic radioligand, [3H]QNB. Particulate fraction binding sites exhibited half maximal binding at approximately 0.3 nM [3H]QNB and reached a maximal binding capacity of 820 fmoles/mg of protein at 3 nM [3H]QNB. Muscarinic cholinergic antagonists and agonists competed with [3H]QNB when incubated concurrently with the tissue, showing relative potencies expected of these agents when binding to muscarinic cholinergic receptors. Particulate fractions prepared from cultured bovine corneal endothelium exhibited qualitatively similar [3H]QNB binding characteristics, but maximal binding capacity was only about one-fifth of its fresh-tissue counterpart. Intact cultured cells showed 3-fold more specific [3H]QNB binding than did their particulate fractions. Incubation of intact corneal endothelial cells with muscarinic cholinergic agonists such as carbachol stimulated cyclic [3H]GMP 3-fold from endogenous [3H]GTP within 1 min of incubation. The effect diminished rapidly and returned to control levels within 8 min. Carbachol stimulation of cyclic [3H]GMP was concentration-dependent, reaching half maximal stimulation at 1 microM. Atropine was a potent, competitive inhibitor of carbachol stimulation of cyclic [3H]GMP in endothelial cultures, requiring only 1 microM to completely block the carbachol response. These experiments demonstrate the existence of muscarinic cholinergic receptors in bovine corneal endothelium and their control of cyclic GMP levels in this tissue.

Cigarette smoking among Beijing (Peking) high schoolers.

PIP: Students smoking at 2 key middle schools and 6 ordinary middle schools in Beijing, China were surveyed to determine the form in which high schoolers start smoking, and how long they have smoked. The patterns and causes of smoking were analyzed. The investigation was made from March to May 1981. 430 boys and 423 girls were selected from key schools and 1396 boys and 1394 girls from ordinary schools. Ages ranged from 13-17. All subjects completed a questionnaire in the classroom. To increase the reliability of the survey, names were kept secret. The students surveyed were classified into groups according to their smoking status: current regular smokers; occasional smokers; ex-smokers; and nonsmokers. In the key schools, there were only 4 smokers among 430 students, accounting for 0.9% of the total. Of these, 1 was Senior Class 2, 2 Senior Class 1, and 1 Junior Class 3. In the ordinary schools, 10% were smokers. 19.7% of the boys and 0.4% of the girls smoked. Among the boys, the higher the class, the more numerous the smokers. The smoking rate was 8.2% in the 1st year of junior high but rose to 34% in senior class 2; the difference was statistically significant. "Special occasion" smokers were few, only 5.5% of the students. In junior class, the number of regular smokers was about 2/3 to 4/5 that of occasional smokers. And by senior class 2 the number of regular smokers had risen to about 3 times that of the occasional smokers. In every grade, there were students who had given up smoking. Most had been occasional smokers. Some high schoolers had started to smoke in primary school and had a 4-5 year history of smoking, but most had only picked up smoking for 1-2 years after announcement of the Rules for High School Students prohibiting smoking. There were 3 smoking patterns: smoking without inhaling; inhaling the smoke deeply into the lungs, then expiring through the mouth or nostrils; and combining patterns 1 and 2. Of 265 students, 128 used the 1st pattern, 121 the second, and only 16 combined both patterns. The most common cause of smoking was curiosity. Next was lack of high ideals and dedication followed by boredom. The smoking rate in students whose parents were smokers was significantly higher than in those whose parents were nonsmokers. The difference was significant among all classes except junior class 1. To reduce smoking among high school students, it is necessary to go beyond the existing rules which prohibit smoking and emphasize political and ideologic education. Students must be taught about the dangers of smoking as early as possible, preferably in primary schools.^ieng