Defect structures in nematic liquid crystals around charged particles.

We numerically study the orientation deformations in nematic liquid crystals around charged particles. We set up a Ginzburg-Landau theory with inhomogeneous electric field. If the dielectric anisotropy epsilon 1 is positive, Saturn-ring defects are formed around the particles. For epsilon 1< 0 , novel "ansa" defects appear, which are disclination lines with their ends on the particle surface. We find unique defect structures around two charged particles. To lower the free energy, oppositely charged particle pairs tend to be aligned in the parallel direction for epsilon 1> 0 and in the perpendicular plane for epsilon 1< 0 with respect to the background director. For identically charged pairs the preferred directions for epsilon 1> 0 and epsilon 1< 0 are exchanged. We also examine competition between the charge-induced anchoring and the short-range anchoring. If the short-range anchoring is sufficiently strong, it can be effective in the vicinity of the surface, while the director orientation is governed by the long-range electrostatic interaction far from the surface.

Mutations of carnitine palmitoyltransferase II (CPT II) in Japanese patients with CPT II deficiency.

Carnitine palmitoyltransferase II (CPT II) deficiency is an inherited disorder involving beta-oxidation of long-chain fatty acids. CPT II deficiency is a wide-spectrum disorder that includes a lethal neonatal form, an infantile form, and an adult-onset form. However, the ethnic characteristics and the relationship between genotype and clinical manifestation are not well understood. We investigated three non-consanguineous Japanese patients with CPT II deficiency and examined cell lines from 4 unrelated patients and 50 healthy donors. The CPT 2 gene was typed by direct DNA sequencing of polymerase chain reaction-amplified gene products. Case 1 (infantile form) was heterozygous for a phenylalanine to tyrosine substitution at position 383 (p.F383Y) and a novel valine to leucine substitution at 605 (p.V605L). Cases 2, 4, and 5 (infantile form) and case 3 (adult-onset form) were heterozygous for a single mutation at F383Y. Case 6 (adult-onset form) was compound heterozygous at the CPT 2 locus, with deletion of cytosine and thymine at residue 408, resulting in a stop signal at 420 (p.Y408fsX420), and an arginine to cysteine substitution at position 631 (p.R631C). Case 7 (adult-onset form) was homozygous for the p.F383Y mutation. In conclusion, we identified p.F383Y mutations in six of seven patients with CPT II deficiency and two novel variants of the coding gene: p.Y408fsX420 and p.V605L. These mutations differ from those in Caucasian patients, who commonly harbor p.S113L, p.P50H, and p.Q413fsX449 mutations; therefore, our data and those of other Japanese groups suggest that the p.F383Y mutation is significant in Japanese patients with CPT II deficiency.

Unexpected monophyletic origin of Ephoron shigae unisexual reproduction strains and their rapid expansion across Japan.

The burrowing polymitarcyid mayfly Ephoron shigae is distributed across Japan, Korea, northeast China and far east Russia. Some populations are bisexual, and others are unisexual, i.e. geographically parthenogenetic throughout Japan. In general, parthenogenetic organisms are often found in harsh environments, such as at high latitudes and altitudes, in xeric as opposed to mesic conditions, in isolated habitats such as islands and island-like areas, and at the peripheral regions of the taxon's range. In E. shigae, however, the distributions of bisexual and unisexual populations overlap broadly in their respective geographical ranges. In the analysis of mitochondrial 16S rRNA and COI, we revealed that unisexual populations were of monophyletic origin and recently differentiated somewhere in western Japan. In the nuclear DNA EFI-α analysis, parthenogenetic strains had two genotypes, i.e. the heterozygous genotype of E1/E3 and the homozygous genotype of E1/E1 or E3/E3, while specimens of bisexual lineage had 20 genotypes. These results are consistent with an automixis mode of reproduction for the parthenogenetic strains, and also support the monophyletic origin of the parthenogenetic strains. Furthermore, there would be no gene flow between the specimens of the bisexual lineage and those of the parthenogenetic strain.

Pharmacokinetic model for in vivo/in vitro correlation of intravitreal drug delivery.

A pharmacokinetic model of intravitreal drug delivery has been developed for describing the elimination and distribution of ocular drugs in the posterior segments of the eye. The model, based on Fick's second law of diffusion, assumes the cylindrical vitreous body with three major pathways for elimination: the posterior aqueous chamber, the retina/choroids/sclera (RCS) membrane and the lens posterior capsule. The model parameters such as the diffusion coefficient and the partition coefficient of the drug in the vitreous body and its surrounding tissues, the posterior lens capsule and the retina/choroids/sclera membrane, can be determined from in vitro membrane penetration experiments using respective rabbit tissues. The time course of in vivo mean concentration of the drug in the rabbit vitreous body following intravitreal drug delivery well agreed with the profile calculated from the present pharmacokinetic model for both membrane-controlled polymeric devices and biodegradable rod-matrix systems. The pharmacokinetic model suggests that the major route of elimination of drug molecules released from the vitreous implant is through the posterior aqueous humor because of the absence of a barrier membrane. However, the elimination through the RCS membrane cannot be overlooked because of the large diffusion area of the RCS membrane. The vitreous body concentration of the drug released from biodegradable vitreous implants can be predicted from the in vivo release rate-time profile by the present pharmacokinetic model.

A method for predicting steady-state rate of skin penetration in vivo.

A simple in vivo method was proposed for predicting the steady-state rate of penetration of drugs across the stratum corneum. Both the diffusion coefficient and the partition coefficient in the stratum corneum can be determined by the amounts of drug in the stratum corneum at two time intervals under transient conditions after transdermal drug application. The amount of drug entering the stratum corneum is determined by 20 strippings with an adhesive tape. The steady-state rate of penetration was then calculated for the thickness of the stratum corneum and the concentration of the donor solution. The steady-state rates of penetration of ascorbic acid and estradiol across hairless mouse skin were evaluated from this in vivo approach and compared with those obtained from in vitro penetration experiment using excised hairless mouse skin. The data confirmed that the proposed in vivo method can predict the steady-state rate of penetration of these drugs across the stratum corneum in normal skin.

Involvement of growth hormone (GH)-releasing factor in GH secretion induced by serotoninergic mechanisms in conscious rats.

A possible role of hypothalamic GH-releasing factor (GRF) in GH secretion induced by serotoninergic stimuli was investigated in conscious freely moving rats with a passive immunization technique. Either iv injection of 5-hydroxy-L-tryptophan (1 mg/100 g BW) or intracerebroventricular injection of serotonin (10 micrograms/rat) caused an increase in plasma GH in rats treated with normal rabbit serum (0.5 ml/rat, iv) 30 min previously. Injection of rabbit antiserum specific for rat GRF (0.5 ml/rat, iv, 30 min previously) blunted the plasma GH increase induced by 5-hydroxy-L-tryptophan or serotonin in these animals. These results suggest that GH secretion induced by serotoninergic mechanisms is mediated, at least in part, by hypothalamic GRF in the rat.

Stimulation of growth hormone secretion by central administration of atrial natriuretic polypeptide in the rat.

Intracerebroventricular (icv) injection of alpha-human atrial natriuretic polypeptide (alpha hANP) or alpha-rat ANP (0.6 and 3 nmol/rat) elicited an increase in plasma GH levels both in conscious freely moving rats and in urethane-anesthetized rats when given at the trough of spontaneous GH secretion. Antiserum specific for rat GRF did not affect the plasma GH increase induced by icv injection of alpha hANP. Intracerebroventricular injection of alpha ANP (3 nmol/rat) failed to stimulate GH secretion in conscious rats pretreated with cysteamine (30 mg/100 g BW, sc), a depletor of somatostatin (SRIF), and in conscious rats during constant iv infusion of SRIF (55 ng/ml). GH release induced by iv injection of synthetic rat GRF (200 ng/100 g BW) was exaggerated by alpha hANP (3 nmol/rat, icv) in conscious rats. These results suggest that central ANP stimulates pituitary GH secretion possibly by inhibiting SRIF release from the hypothalamus in the rat.

Involvement of growth hormone-releasing factor in growth hormone secretion induced by gamma-aminobutyric acid in conscious rats.

Intracerebroventricular (icv) injection of gamma-aminobutyric acid (GABA) (10 mumol/rat) resulted in an increase in plasma GH in conscious freely moving rats pretreated with normal rabbit serum (0.5 ml/rat, iv). Rabbit antiserum specific for rat GH-releasing factor (GRF) (0.5 ml/rat, iv) abolished GH release induced by GABA in these animals. Rabbit anti-rat GRF serum also blunted GH release induced by a Met5-enkephalin analog, FK33-824 (10 micrograms/100 g BW, iv) in conscious rats. Considering our previous findings that rat GH release induced by FK33-824 was blunted by GABA antagonists (Endocrinology 103:1033, 1981), these results suggest that GH secretion induced by opioid peptides via GABAergic mechanisms is mediated, at least in part, by hypothalamic GRF in the rat.

Stimulation by leumorphin of prolactin secretion from the pituitary in rats.

The effect of leumorphin (LM), one of big leu-enkephalins derived from preproenkephalin B, on PRL secretion was studied in the rat in vivo and in vitro. Intracerebroventricular injection of synthetic porcine LM (0.06-6 nmol/rat) caused a dose-related increase in plasma PRL levels in urethane-anesthetized male rats and in conscious freely moving rats. Intravenous injection of LM (3 nmol/100 g BW) also raised plasma PRL levels in these animals. The plasma PRL response to intracerebroventricular LM (0.6 nmol/rat) was blunted by naloxone (125 micrograms/100 g BW, iv). The stimulating effect of LM on PRL release was the most potent among the peptides derived from preproenkephalin B. In in vitro studies, PRL release from superfused anterior pituitary cells was stimulated in a dose-related manner by LM (10(-9)-10(-6) M), and the effect was blunted by naloxone (10(-5) M). These results suggest that LM has a potent stimulating effect on PRL secretion from the pituitary in the rat by acting, at least in part, directly at the pituitary through an opiate receptor.

Penetration and binding of aldose-reductase inhibitors in the lens.

Diffusion, partitioning, and binding of two aldose-reductase inhibitors (ARI) in the rat lens were investigated under in vitro conditions. A lipophilic ARI (CT-112) and a hydrophilic ARI (AD-5467) were used as model drugs. Under lens-uptake conditions, the drug concentration in the lens increased rapidly during the initial 10 hr and reached steady state (equilibrium) after 24 hr. Despite the equilibrium concentration of CT-112 in the lens which is approximately three times of that of AD-5467, the inhibition rate of CT-112 against the accumulation of sugar alcohols was appreciably lower than that for AD-5467. The equilibrium concentration in the lens after the penetration/binding experiment also suggested that AD-5467 may interact with the target sites of enzymes more than that for CT-112. The time course of the lens concentration during the uptake and desorption experiments was well described by a diffusion/binding model assuming a Langmuir binding. The diffusion coefficient, the partition coefficient, and binding parameters in the rat lens were determined for the two ARIs.

Possible involvement of endothelin-1 in cardioprotective effects of benidipine.

Benidipine hydrochloride has been developed as an antagonist for the L-type calcium channel and is used as an anti-hypertensive drug. But recent studies have reported that benidipine exerts not only antihypertensive actions but also anti-hypertrophic actions on cardiac muscles. Endothelin-1 (ET-1), one of the endogenous pathological humoral factors of cardiovascular diseases such as hypertension and heart failure, has a strong vasoconstrictive action and could induce hypertension and cardiac hypertrophy. So, it is a matter of great interest whether or not calcium antagonists can decrease cardiac hypertrophy induced by the pathological vasoactive substances such as ET-1. Thus, the present study was designed to elucidate the effects of benidipine on cardiac hypertrophy, and particularly on the interaction with ET-1, using neonatal rat cardiac myocytes (MCs) and cardiac non-myocytes (NMCs) culture systems. Cells were cultured with or without ET-1, benidipine, and nifedipine and the effects of calcium antagonists on cardiac hypertrophy were evaluated by incorporations of [3H]-leucine and [3H]-thymidine into MCs and/or NMCs. Benidipine significantly decreased the ET-1-induced increase of [3H]-leucine and [3H]-thymidine uptake into cardiac MCs and NMCs, whereas no significant effects of nifedipine were observed. Furthermore, benidipine (10(-8)M) attenuated ET-1 secretions from NMCs. In summary, benidipine at least partially decreased the cardiac hypertrophy induced by paracrine mechanisms through its attenuation of ET-1 secretions from NMCs. Benidipine could thus be a useful tool for preventing cardiac hypertrophy due to hypertension.

Further evidence that peptide histidine isoleucine (PHI) may function as a prolactin releasing factor in rats.

Intraventricular administration of peptide histidine isoleucine (PHI) (200 ng, 1, 5 and 10 micrograms/rat) resulted in a significant and dose-related increase in plasma prolactin (PRL) levels in urethane-anesthetized rats and in conscious rats with intraatrial and intraventricular catheters. Intravenous injection of PHI (10 micrograms/rat) also raised plasma PRL levels in these animals. In in vitro studies, PRL release from superfused rat anterior pituitary cells was stimulated by PHI (10(-9), 10(-8) and 10(-7) M) in a dose-related manner. The stimulating effect of PHI (10(-7)M) on PRL release in vitro was as potent as that of vasoactive intestinal polypeptide (VIP) (10(-7) M) and was observed even in the presence of dopamine (10(-7) M). These results suggest that PHI plays a stimulating role in regulating PRL secretion by acting, at least in part, directly on the pituitary in the rat.

Inhibition by antiserum to vasoactive intestinal polypeptide (VIP) of prolactin secretion induced by serotonin in the rat.

Intraventricular (i.c.v.) injection of serotonin (5HT) or intravenous (i.v.) injection of 5-hydroxytryptophan (5HTP), a precursor of 5HT, raised plasma prolactin (PRL) levels in urethane-anesthetized rats pretreated with normal rabbit serum. When the animals were injected i.c.v. or i.v. with specific anti-VIP rabbit serum, the plasma PRL responses to 5HT and 5HTP were blunted. These findings suggest that hypothalamic VIP is involved, at least in part, in PRL secretion induced by central serotonergic stimulation in the rat.

Stimulation by serotonin of immunoreactive peptide histidine isoleucine release from rat hypothalamus in vitro.

The effect of serotonin (5-HT) on the release of peptide histidine isoleucine-like immunoreactivity (PHI-LI) from rat hypothalamus was investigated in vitro with a perifusion system. A high potassium concentration (56 mM) stimulated PHI-LI release in a calcium-dependent manner. PHI-LI release was dose-relatedly stimulated by 5-HT (10(-6), 10(-5) and 10(-4) M). PHI-LI release induced by 5-HT (10(-5) M) was abolished by cyproheptadine (10(-4) M), a 5-HT antagonist. These results suggest that 5-HT has a stimulating effect on PHI release from the hypothalamus.

Corticotropin-releasing factor (CRF) stimulates 45Ca2+ uptake in the mouse corticotroph cell line AtT-20.

Corticotropin-releasing factor (CRF) stimulates adrenocorticotropin (ACTH) release via the adenylate cyclase/cAMP-dependent protein kinase system. Because calcium is necessary for receptor-mediated release of ACTH, we have examined the effect of CRF on 45Ca2+ uptake in a corticotroph cell line model, AtT-20. Treatment of AtT-20 cells with CRF (10(-9)-10(-6) M) resulted in dose- and time-dependent increases in 45Ca2+ uptake, up to 2.2-fold above control values. The effect was statistically significant at 1 min and persisted for at least 10 min. Treatment with forskolin (1-30 microM), 8-Br-cAMP (0.5 mM), cholera toxin (CT, 100 ng/ml) and K+ (20 mM) also increased cell-associated 45Ca2+. The effect of K+ was completely blocked by nifedipine (100 microM), whereas the effects of CRF (10(-8) M) were only partially inhibited by this calcium channel antagonist. These data suggested a role of voltage-dependent calcium channels in 45Ca2+ uptake. Short term pretreatment (1-2 h) of AtT-20 cells with CRF (10(-8) M) significantly desensitized both CRF-stimulated cAMP accumulation and ACTH release, but did not attenuate CRF-stimulated 45Ca2+ uptake. Pretreatment with CRF (10(-8) M) for 4 h did not alter CT- or forskolin-stimulated cAMP accumulation and ACTH release. This suggests that the molecular mechanisms of desensitization are proximal to adenylate cyclase. Conversely, long term pretreatment (24 h) of AtT-20 cells with CRF (10(-8) M) induced significant desensitization of CRF-stimulated 45Ca2+ uptake. These results indicate that CRF stimulates calcium uptake in AtT-20 cells via cAMP-dependent and cAMP-independent mechanisms, and that the cellular mechanisms involved in desensitization of cAMP accumulation and ACTH release and those involved in desensitization of calcium uptake are qualitatively different.

The effects of sarpogrelate on cardiomyocyte hypertrophy.

Sarpogrelate was developed as an antiplatelet agent antagonizing 5-hydroxytryptamine (5-HT) receptors. It had been reported that 5-HT receptors were expressed in cardiovascular system, and that sarpogrelate had antihypertrophic effects in vascular smooth muscle cells. Cardiac hypertrophy is a major problem in cardiac diseases, so the present study was designed to elucidate the effects of sarpogrelate on cardiac hypertrophy. Cultured rat cardiomyocytes (MCs) and cardiac nonmyocytes (NMCs) were prepared by Percoll gradient and adhesion method and MCs were incubated with (MCs/NMCs) or without NMCs. As an index of protein synthesis of MCs, [3H]-leucine uptake into MCs and MCs/NMCs was measured. Sarpogrelate decreased [3H]-leucine uptake into MCs (maximum 62.6+/-20.6% of control at 10(-4)M, p<0.05 vs. control). Sarpogrelate also significantly attenuated angiotensin-II- and endothelin-1-induced [3H]-leucine uptake. These results indicated that sarpogrelate might have antihypertrophic effects and could be a useful aid for cardiovascular disease.

Central effects of DN1417, a novel TRH analog, on plasma glucose and catecholamines in conscious rats.

Intracerebroventricular (icv) injection of DN1417 (0.3, 3 and 30 nmol/rat), a TRH analog, resulted in a dose-related increase in plasma glucose, epinephrine and norepinephrine levels in conscious male rats. The effects of DN1417 were more potent and longer-lasting than those of TRH on a molar basis. Intravenous injection of DN1417 (30 nmol/rat) did not change plasma glucose, epinephrine and norepinephrine levels. Pretreatment with hexamethonium (1.5 mg/100 g body wt, iv, 2 min before) inhibited plasma glucose, epinephrine and norepinephrine responses to DN1417 (3 nmol/rat, icv). DN1417 did not change plasma glucose, epinephrine and norepinephrine levels in rats after total adrenalectomy. In the animals pretreated with cysteamine (30 mg/100 g body wt, sc, 4 h before), basal plasma glucose, epinephrine and norepinephrine levels were raised, and exaggerated responses of plasma glucose, epinephrine and norepinephrine to DN1417 (3 nmol/rat, icv) were obtained. These results indicate that DN1417 has a potent and long-lasting effect in the central nervous system in stimulating the secretion of catecholamines through the autonomic nervous system, which is associated with an elevation of plasma glucose and that endogenous hypothalamic somatostatin may inhibit the action of DN1417.

A pharmacokinetic model of intravitreal delivery of ganciclovir.

A pharmacokinetic model of intravitreal drug delivery was developed for describing the elimination and distribution of ganciclovir in the eye following intravitreous polymeric delivery. The model was based on Fick's second law of diffusion and assumed a cylindrical vitreous body. The model parameters such as the diffusion coefficient and the partition coefficient of the drug in the vitreous body and its surrounding tissues were determined from in vitro experiments using rabbit tissues. The time course of in vivo mean concentration of ganciclovir in the rabbit vitreous body agreed well with the profile calculated from the present pharmacokinetic model for both membrane-controlled polymeric devices and biodegradable rod-matrix systems. The clinical vitreous concentration following implantation of the membrane-controlled delivery system was the same order of magnitude but approximately four times lower than that predicted from the present model. This may indicate the metabolism of ganciclovir and/or the facilitated transport across the retina/choroid membrane in the human eye.

Random brick model for drug transport across stratum corneum.

A model for drug permeation across the stratum corneum is developed to predict the effect of the physical configuration and chemical composition of the stratum corneum on the skin permeability of drugs. The experimental data of percutaneous absorption across different skin specimens are well interpreted by the present model.

Intrinsic release rate from matrix-type drug delivery systems.

A simple method was presented for evaluation of the intrinsic release rate from a matrix-type drug delivery system. The effect of the hydrodynamic diffusion boundary layer on the rate of drug release was corrected to obtain the release profile without any diffusion boundary layer effect. The experimental release data was logically explained in terms of the intrinsic rate of release evaluated from the present method.