Biomedical subjects
Publications and source records attributed to K Kurihara.
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Dahl-Iwai salt-sensitive (S) and salt-resistant (R) rat strains were established as inbred strains at Brookhaven National Laboratory, NY, and were introduced into Eisai Co., Ltd., Japan, and designated DIS/Eis and DIR/Eis. To examine whether there are different allele distributions among the substrains of inbred Dahl S and R rats, we determined biochemical and immunological alleles of DIS/Eis and DIR/Eis, and SS/Sea and SR/Sea, which were derived from SS/Jr and SR/Jr, which were developed by Rapp and Dene. Several differences of allele distribution were observed, indicating that the substrains have different genetic backgrounds. The phenotypic differences between the substrains, such as the severity of the hypertension induced, could be ascribed to the different genetic backgrounds.
Multicentric breast carcinomas not diagnosed clinically, were examined by serial step-cut sectioning of the whole breast, and multicentric carcinoma cases were compared with single carcinoma cases with regard to histological and clinicopathological findings. In 7(3.7%) out of 187 surgically resected breasts, latent carcinomas apart from the main carcinoma were noted. The size of the latent carcinoma varied from 0.2 to 5 cm in diameter. The histological type was noninvasive ductal carcinoma in six cases and invasive ductal carcinoma in one case. When the main carcinoma was small in size (less than 2.5 cm in diameter), and showed papillotubular carcinoma as the histological type or had estrogen receptor (ER) by the dextran-coated charcoal (DCC) method, the incidence of latent carcinoma was high. In 5 of 6 cases with latent carcinoma examined by immunohistochemistry, latent carcinomas showed expression of ER. Concerning Ki-67, proliferating cell nuclear antigen (PCNA) and p53 protein, there was no significant difference between the main and latent carcinomas, as well as with other clinicopathological factors.
The responses of single bullfrog olfactory neurons to various odorants were measured with the whole-cell patch clamp which offers direct information on cellular events and with the ciliary recording technique to obtain stable quantitative data from many neurons. A large portion of single olfactory neurons (about 64% and 79% in the whole-cell recording and in the ciliary recording, respectively) responded to many odorants with quite diverse molecular structures, including both odorants previously indicated to be cAMP-dependent (increasing) and independent odorants. One odorant elicited a response in many cells; e.g. hedione and citralva elicited the response in 100% and 92% of total neurons examined with the ciliary recording technique. To confirm that a single neuron carries different receptors or transduction pathways, the cross-adaptation technique was applied to single neurons. Application of hedione to a single neuron after desensitization of the current in response to lyral or citralva induced an inward current with a similar magnitude to that applied alone. It was suggested that most single olfactory neurons carry multiple receptors and at least dual transduction pathways.
During metamorphic stages (stages XX-XXV), the first appearance of nerve fibers containing substance P (SP), calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP), galanin (GAL), and neuropeptide Y (NPY) in the bullfrog taste organs was different for each substance. CGRP fibers appeared first in association with the immature taste organs at stage XX. Up to stage XXV, the taste organs, epithelial disks, are close to their adult form, and SP, VIP and GAL fibers appeared within them. Throughout these stages, NPY fibers were absent, and no taste cells had immunoreactivity of the five neuropeptides. The present findings and previous physiological studies suggest that the immature taste organs in metamorphic stages already function as chemical and mechanical receptors and that these sensory mechanisms are under the control of peptidergic innervation.
An application of double-immunolabelling in combination with a multiple dye filter system demonstrated new findings regarding the distribution pattern of peptidergic fibers in the carotid labyrinth to addition to our previous findings shown by the individual filter system. In high magnification images of about 10% of the yellowish fibers which represent the coexistence of two neuropeptides, there was a definite difference in localization between the fluorescence originating from rhodamine (substance P fibers) and from FITC (vasoactive intestinal polypeptide and neuropeptide Y fibers), but it was clear that they are intertwined within a single nerve bundle. This combination method was able to discriminate two different peptidergic fibers which run side by side. The coexistence suggested previously by the individual filter system may actually be due to the phenomenon described above. This means that it is necessary to apply the multiple dye filter system for reliable evidence of coexistence of different two substances in a single nerve fiber.
Ciliary neurotrophic factor (CNTF) is a neuropoietic cytokine which has various functions, such as survival promoting effect on both peripheral and central neurons, promotion of cholinergic differentiation, and participation in differentiation of Type-2 astrocytes (reviewed in ref. [30]). However, the regulatory mechanism of the CNTF expression is largely unknown. In this study, we analyzed the effects of phorbol 12-myristate 13-acetate (PMA), an activator of PKC, on the expression of CNTF-mRNA in cultured astrocytes from neonatal rat olfactory bulb. PMA induced a transient decrease of CNTF-mRNA levels which was followed by a persistent increase of the mRNA up to 4-fold of the control level at 24 h after the addition of the compound. Both the PMA-induced decrease and increase of the CNTF-mRNA levels were canceled by treatment with cycloheximide, an inhibitor of protein synthesis, suggesting that protein synthesis-dependent mechanisms participate in both the PMA-induced decrease and increase of CNTF-mRNA levels. On the other hand, PMA induced expressions of mRNAs of several subunit members of the AP-1 complex, such as c-fos, c-jun and jun-B. Furthermore, dexamethasone, a synthetic glucocorticoid which is known to inhibit the AP-1 complex-mediated transcription [14,27,36], canceled the PMA-induced decrease of the CNTF-mRNA levels. These results suggested that the AP-1 complex participates in the regulatory mechanism of the CNTF expression in the cultured astrocytes treated with PMA.
In previous studies (Katsuragi and Kurihara (1993) Nature 365,213--214; Katsuragi et al. (1995) Pharm. Res. 12,658--662) we showed that a lipoprotein composed of phosphatidic acid (PA) and beta-lactoglobulin (LG) selectively suppressed the taste responses to bitter substances without affecting those to other taste stimuli in the frog and man, while complexes composed of other lipids except for phosphatidylserine and LG had little inhibitory activity. In the present study, we found that the lipoproteins having inhibitory activity are adsorbed on the frog tongue surface, while those having no inhibitory activity are not adsorbed. We also examined adsorption of the lipoproteins on model lipid membranes coated on a quartz-crystal microbalance by measuring changes in its frequency. The lipoproteins having inhibitory activity were well adsorbed on the hydrophobic lipid membranes, while the lipoproteins having no inhibitory activity were little adsorbed on the membranes. It seems that receptor sites for bitter substances on the taste cell membranes are hydrophobic and those for other taste stimuli such as salts, acids and sugars are hydrophilic. Hence, the binding of PA-LG to hydrophobic sites of the receptor membranes will lead to selective inhibition of bitterness.
The development of a specific inhibitor for bitter taste has been widely required in the fields of taste physiology and pharmaceutical sciences, but no inhibitor has been available. We found that lipoproteins, PA-LG composed of phosphatidic acid (PA) and beta-lactoglobulin (LG) and PA-LA composed of PA and alpha-lactalbumin (LA) reversibly suppressed the responses of the frog glossopharyngeal nerve to the bitter substances. The frog tongue was treated with PA-LG solution for 10 min and then stimulated by a stimulus dissolved in water. The responses to the bitter substances such as quinine hydrochloride, papaverine hydrochloride, caffeine and L-leucine were completely suppressed by PA-LG, while those to the salt type bitter substances such as CsCl, MgCl2 and tetraethylammonium chloride were not suppressed. The responses to NaCl, galactose, acetic acid and L-alanine were unchanged or only slightly increased. The results suggested that binding of PA-LG to the hydrophobic region of the receptor membranes leads to suppression of the responses to the bitter substances. It was pointed out that PA-LG is useful not only for elucidating the receptor mechanisms of bitter substances, but also can be safely used to mask the bitter taste of foods and drugs, since PA, LG and LA are prepared from foods (soybean and milk).
Turtle vomeronasal receptor neurons in slice preparations were studied using the patch-clamp technique in the whole-cell and cell-attached configurations. The mean resting potential was -48, and the response to an injected current step consisted of either a single spike or a train of spikes. An injected current of 3-30 pA was required to depolarize the neuron to spike threshold near -50 mV. Voltage-clamped vomeronasal receptor neurons displayed transient inward currents followed by sustained outward currents in response to depolarizing voltage steps. In cell-attached recordings, 10 microM forskolin added to the bath caused a transient increase of spike rate. Intracellular application of cAMP evoked ann inward current in a dose-dependent manner from the neurons voltage clamped at -70 mV; 0.1 mM cAMP was sufficient to elicit an inward current in the neurons. The magnitude of the response to cAMP reached a plateau at 1 mM with an average peak amplitude of 176 +/- 34 pA. Intracellular application of 1 mM cGMP also evoked an inward current with an average peak amplitude of 227 +/- 61 pA. The reversal potentials of the induced components were estimated to be 10 +/- 7 mV for cAMP and -4 +/- 16 mV for cGMP. The reversal potential of the cAMP-induced current in external Cl(-)-free solution was similar to that in normal Ringer's solution, suggesting that Cl- current is not significantly involved in the current. The present results represent the first evidence of cyclic nucleotide-activated conductance in the vomeronasal receptor membranes.
The ability of the turtle olfactory system to discriminate between various odorants that increase levels of adenosine 3',5'-cyclic monophosphate (cAMP) and inositol trisphosphate (IP3) in the olfactory bulb was examined by the cross-adaptation technique and analyzed by multidimensional scaling. The mean values of the degree of discrimination among the IP3-increasing odorants were higher than those among the cAMP-increasing odorants, and were similar to those between cAMP- and IP3-increasing odorants, suggesting that the features of the receptors of cAMP-increasing odorants are different from those which respond to IP3-increasing odorants. Analysis by multidimensional scaling suggested that differences in second messenger pathways are not related to detecting odor quality in the turtle olfactory system.
In the turtle olfactory system, large responses to odorants appeared after application of cAMP of forskolin at high concentrations to the isolated olfactory receptor neurons or the olfactory epithelium, suggesting that a cAMP-independent pathway greatly contributes to the generation of odor responses. We measured the effects of the mucosal Ca2+ and Cl- concentrations and a Ca(2+)-activated Cl(-)-channel blocker, 4-acetamide-4'-(isothiocyano) stilbene-2,2'-disulphonic acid (SITS), upon olfactory bulbar responses to explore the contribution of the Ca(2+)-activated Cl(-)-channels to cAMP-dependent and independent pathways. Elimination of mucosal Ca2+ by addition of 2 mM EGTA to the stimulating solution partially inhibited the cAMP-independent responses to 0.1 mM citralva but did not affect those to 0.1 mM lilial or 0.1 mM l-carvone. Substitution of mucosal Cl- with gluconate slightly enhanced the total response to 0.1 mM citralva but practically did not affect the responses to other odorants tested. SITS (2 mM) partially inhibited the responses to 50 microM forskolin and 0.1 mM citralva but did not affect the cAMP-independent response to 0.1 mM citralva in Ca(2+)-free Ringer's solution. These results suggested that the Ca(2+)-activated Cl- channels do not contribute to the generation of the cAMP-independent responses, but that they partially contribute to the generation of the cAMP-dependent responses in the turtle.
The cells of the granular convoluted tubule (GCT) of the rodent submandibular gland (SMG) are under complex developmental and multihormonal regulation. Recent findings indicate that GCT cells also synthesize transforming growth factor alpha (TGF-alpha), hepatocyte growth factor, erythroid differentiation factor, endothelin, and insulin-like growth factor, as well as several novel androgen-dependent proteins of unknown function. The GCTs of hypophysectomized mice provide a convenient model to study multihormonal regulation of gene expression. The GCT system of the rodent SMG also is a fruitful model for study of hormone receptors.
The [(3)H]inositol incorporation into the membrane fraction of A-431 human epidermoid carcinoma cells was markedly increased by stimulation of the cells with either epidermal growth factor (EGF), ATP, bradykinin, or a calcium ionophore A23187 in the presence of 1 mM extracellular calcium ions; most incorporated [(3)H]inositol was found to have accumulated as phosphatidylinositol (PI). The EGF- and ATP-stimulated PI synthesis was inhibited by two protein kinase C inhibitors, staurosporine and 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7), and an intracellular calcium chelator, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester (BAPTA/AM), but not by the calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7). Pretreatment of cells with pertussis toxin (IAP, islet-activating protein) inhibited the PI synthesis, [Ca(2+)]i elevation, and inositol trisphosphate (IP(3)) production by ATP, suggesting that the phospholipase C(PLC) system coupled with IAP-sensitive G protein is involved in the ATP-stimulated PI synthesis. On the other hand, the ATP stimulation increased the release of [(3)H]choline and [(32)P)phosphatidic acid (PA) from radiolabeled cells, and such release was not inhibited by IAP. In the presence of n-butyl alcohol, which prevents the production of PA by generation of phosphatidylbutanol, the ATP-stimulated PI synthesis was reduced. Because n-butyl alcohol did not inhibit IP(3) production and [Ca(2+)]i elevation, this fact suggests that the lAP-insensitive PLD system is involved in the ATP-stimulated PI synthesis. In A-431 cells, the stimulation of P(2)-purinergic receptors appears to activate the IAP-sensitive PLC system and IAP-insensitive PLD system, both of which are essential for the stimulation of PI synthesis. The present results imply the general prospect that ligand stimulation, which mobilizes second messengers and consumes their precursors, simultaneously provokes the pathway to synthesize and salvage the second messenger precursors as well.
The effects of thyroid hormone, androgen, glucocorticoid, and mineralocorticoid on Na+,K+-ATPase activity and on levels of its alpha-subunit protein (alpha 1 isoform) in mouse submandibular gland (SMG) were studied by enzyme assay for ouabain-sensitive ATP hydrolysis, by quantitative densitometric scanning of Western blots, and by immunohistochemistry. To define the specific regulatory effects of various pituitary-dependent hormones on expression of Na+,K+-ATPase in the SMG, we treated hypophysectomized (hypox) male mice with triiodo-L-thyronine (T3), 5 alpha-dihydrotestosterone (DHT), dexamethasone (Dex), and aldosterone (Ald), injected singly or in combination. Na+,K+-ATPase was confined to the duct system of the SMG. In intact mice there was a gender difference in SMG Na+,K+-ATPase, with levels of the enzyme's activity and of its alpha 1-subunit being less in the glands of males. In males, hypophysectomy caused a rise in levels of Na+,K+-ATPase activity and in levels of the alpha 1-subunit protein of this enzyme, and in intensity of immunocytochemical staining for this subunit but there were no such changes in the SMG of hypox females. Changes caused by hormonal replacement to hypox males in Na+,K-ATPase activity, levels of its alpha 1-subunit, or the intensity of immunocytochemical staining for this subunit were complex. Ald had no effect. T3 or dexamethasone, given alone, induced Na+,K+-ATPase activity above control values (hypox males) and increased levels of its alpha 1-subunit protein and immunohistochemical staining for this subunit. By contrast, DHT did not cause a decline in any of these parameters. However, when treatment with T3 was combined with administration of Dex or DHT, enzymatic activity of Na+,K+-ATPase decreased but levels of the alpha 1-subunit protein and immunohistochemical staining for this subunit increased. Therefore, inductions of the alpha 1-subunit of this enzyme are not always correlated with increases in levels of activity of Na+,K+-ATPase, and we propose that both enzymatic and immunochemical analyses are essential for evaluation of hormonal regulation of Na+,K+-ATPase in salivary gland and in other tissues.
Salmonid fishes are able to survive in both fresh water and sea water. Concentrations of NaCl in fresh water and sea water are 0.5 and 493 mmol l-1, respectively, and, hence, salt concentrations in the medium at the olfactory epithelium are greatly changed when the fish migrate between fresh water and sea water. We used the rainbow trout, which is a salmonid fish, to examine the adaptation mechanisms of the olfactory receptors to high concentrations of salts in sea water. Application of sea water to the olfactory epithelium elicited only a very small response in the olfactory nerve, but 500 mmol l-1 NaCl elicited a large response which did not adapt to a spontaneous level with time. It is considered that the olfactory nerve becomes fatigued when the olfactory epithelium is exposed to 500 mmol l-1 NaCl for long periods. We found that the presence of 10 mmol l-1 Ca2+ in sea water inhibited the response to 500 mmol l-1 NaCl, suggesting that Ca2+ in sea water is essential for adaptation of the olfactory receptors to sea water. In the second part of the study, we examined whether the olfactory nerve responses to amino acids, potent stimulants for fish, were altered between fresh water and sea water. The magnitudes of the responses to the six amino acids examined were similar in artificial pond water and artificial sea water, indicating that a large change in NaCl concentration between fresh and sea water does not affect the olfactory nerve responses to amino acids. We used fish reared in fresh water and fish acclimated to sea water and obtained similar results. It was concluded that the tolerance of the olfactory receptors for large changes in osmotic pressure is not acquired while fish are maintained in fresh or sea water, but that the receptors of these euryhaline fish naturally provide the tolerance.
We describe a 60-year-old man with a history of Tolosa-Hunt syndrome associated with intermittent painful ophthalmoplegia and a visual disturbance on the left side, who presented with signs and symptoms of severe hypoadrenalism and diabetes insipidus. Magnetic resonance imaging demonstrated enlargement of the hypophysis and infundibulum and left cavernous sinus. An endocrinologic study revealed anterior pituitary dysfunction and diabetes insipidus. The patient underwent a transsphenoidal biopsy which revealed chronic inflammation in the hypophysis, mucosa of the sphenoid sinus, and dura mater. The patient was treated with steroids that decreased the size of the hypophysis and infundibulum, but the symptoms of anterior pituitary insufficiency and diabetes insipidus have persisted. The chronic inflammation of the hypophysis and infundibulum is thought to have spread from the cavernous sinus.