RESUMEN
Three isoforms of the inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3] receptor have been identified. Each receptor isoform has been functionally characterized using many different techniques. Although these receptor isoforms possess high homology, interesting differences in their Ca2+ dependence, Ins(1,4,5)P3 sensitivity and subcellular distribution exist, implying distinct cellular roles. Indeed, interplay among the isoforms might be necessary for a cell to control spatial and temporal aspects of cytosolic Ca2+ signals, which are important for many cellular processes. In this review isoform-specific functions, primarily at the single-channel level, will be highlighted and these properties will be correlated with Ca2+ signals in intact cells.
Asunto(s)
Canales de Calcio/fisiología , Receptores de Superficie Celular/fisiología , Receptores Citoplasmáticos y Nucleares/fisiología , Animales , Canales de Calcio/efectos de los fármacos , Humanos , Receptores de Inositol 1,4,5-Trifosfato , Fosforilación , Receptores de Superficie Celular/química , Receptores de Superficie Celular/genética , Receptores Citoplasmáticos y Nucleares/efectos de los fármacosRESUMEN
Many hormones and neurotransmitters raise intracellular calcium (Ca(2+)) by generating InsP(3) and activating the inositol 1,4, 5-trisphosphate receptor (InsP(3)R). Multiple isoforms with distinct InsP(3) binding properties () have been identified (). The type III InsP(3)R lacks Ca(2+)-dependent inhibition, a property that makes it ideal for signal initiation (). Regulation of the type III InsP(3)R by InsP(3) and ATP was explored in detail using planar lipid bilayers. In comparison to the type I InsP(3)R, the type III InsP(3)R required a higher concentration of InsP(3) to reach maximal channel activity (EC(50) of 3.2 microM versus 0.5 microM for the types III and I InsP(3)R, respectively). However, the type III InsP(3)R did reach a 2.5-fold higher level of activity. Although activation by InsP(3) was isoform-specific, regulation by ATP was similar for both isoforms. In the presence of 2 microM InsP(3), low ATP concentrations (<6 mM) increased the open probability and mean open time. High ATP concentrations (>6 mM) decreased channel activity. These results illustrate the complex nature of type III InsP(3)R regulation. Enhanced channel activity in the presence of high InsP(3) may be important during periods of prolonged stimulation, whereas allosteric modulation by ATP may help to modulate intracellular Ca(2+) signaling.
Asunto(s)
Adenosina Trifosfato/metabolismo , Canales de Calcio/metabolismo , Inositol 1,4,5-Trifosfato/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Adenosina Trifosfato/farmacología , Regulación Alostérica/efectos de los fármacos , Animales , Unión Competitiva/efectos de los fármacos , Canales de Calcio/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Retículo Endoplásmico/efectos de los fármacos , Retículo Endoplásmico/metabolismo , Inositol 1,4,5-Trifosfato/farmacología , Receptores de Inositol 1,4,5-Trifosfato , Insulinoma/metabolismo , Insulinoma/patología , Microsomas/efectos de los fármacos , Microsomas/metabolismo , Isoformas de Proteínas/efectos de los fármacos , Isoformas de Proteínas/metabolismo , Ratas , Receptores Citoplasmáticos y Nucleares/efectos de los fármacos , Especificidad por Sustrato , Células Tumorales CultivadasRESUMEN
The type III inositol 1,4,5-trisphosphate receptor (InsP3R) is an important intracellular calcium (Ca2+) release channel in the pancreatic beta cell. Pancreatic beta cells secrete insulin following a characteristic change in membrane potential that leads to an increase in cytoplasmic Ca2+. Both extracellular Ca2+ and Ca2+ mobilized from InsP3-sensitive stores contribute to this increase. RIN-m5F cells, an insulin-secreting beta cell line, preferentially express the type III InsP3R. These cells have been useful in determining the regulatory properties of the type III InsP3R and the role of this isoform in an intact cell. The type III InsP3R is ideal for signal initiation because high cytoplasmic Ca2+ does not inhibit its activity. Altered insulin secretion, the result of changes in Ca2+ handling by the beta cell, has significant clinical consequences.
Asunto(s)
Canales de Calcio/metabolismo , Señalización del Calcio , Islotes Pancreáticos/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Animales , Calcio/metabolismo , Canales de Calcio/química , Línea Celular , Diabetes Mellitus/metabolismo , Diabetes Mellitus/patología , Conductividad Eléctrica , Humanos , Hipoglucemia/metabolismo , Hipoglucemia/patología , Receptores de Inositol 1,4,5-Trifosfato , Insulina/metabolismo , Secreción de Insulina , Islotes Pancreáticos/patología , Receptores Citoplasmáticos y Nucleares/antagonistas & inhibidores , Receptores Citoplasmáticos y Nucleares/química , SíndromeRESUMEN
The inositol 1,4,5-trisphosphate receptor (InsP3R) is the main calcium(Ca2+) release channel in most tissues. Three isoforms have been identified, but only types I and II InsP3R have been characterized. Here we examine the functional properties of the type III InsP3R because this receptor is restricted to the trigger zone from which Ca2+ waves originate and it has distinctive InsP3-binding properties. We find that type III InsP3R forms Ca2+ channels with single-channel currents that are similar to those of type I InsP3R; however, the open probability of type III InsP3R isoform increases monotonically with increased cytoplasmic Ca2+ concentration, whereas the type I isoform has a bell-shaped dependence on cytoplasmic Ca2+. The properties of type III InsP3R provide positive feedback as Ca2+ is released; the lack of negative feedback allows complete Ca2+ release from intracellular stores. Thus, activation of type III InsP3R in cells that express only this isoform results in a single transient, but global, increase in the concentration of cytosolic Ca2+. The bell-shaped Ca2+-dependence curve of type I InsP3R is ideal for supporting Ca2+ oscillations, whereas the properties of type III InsP3R are better suited to signal initiation.
Asunto(s)
Canales de Calcio/metabolismo , Calcio/metabolismo , Inositol 1,4,5-Trifosfato/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Adenosina Trifosfato/metabolismo , Regulación Alostérica , Animales , Señalización del Calcio , Línea Celular , Citoplasma/metabolismo , Perros , Retículo Endoplásmico/metabolismo , Receptores de Inositol 1,4,5-Trifosfato , RatasRESUMEN
The novel acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor CP-113,818 has been characterized in vitro against ACAT isolated from liver and intestine from a variety of species including human subjects, and in vivo in cholesterol-fed rats, hamsters, rabbits, and two species of nonhuman primates. CP-113,818 is a potent and specific inhibitor of liver and intestinal ACAT with IC50s ranging from 17 to 75 nM. This ACAT inhibitor also prevented the absorption of exogenous radiolabeled cholesterol in hamsters (ED50 = 6 micrograms/kg), rabbits (ED50 1/2 10 micrograms/kg), and cynomolgus and African green monkeys (40 and 26% inhibition at 10 mg/kg, respectively). CP-113,818 effectively prevented the increase in liver cholesterol levels in cholesterol-fed rats, hamsters, and rabbits. In lipoprotein characterization studies in rabbits, CP-113,818 selectively decreased apoB-containing lipoproteins (beta-VLDL, IDL, and LDL) and shifted the distribution of cholesterol from beta-VLDL, IDL, and LDL (96% before treatment to 81% after treatment) to HDL (4% before treatment to 19% after treatment). Finally, in monkeys, CP-113,818 significantly decreased LDL cholesterol by approximately 30% while either increasing HDL cholesterol (cynomolgus monkeys) or not affecting HDL cholesterol (African green monkeys), thereby improving the total plasma cholesterol/HDL ratios. In summary, CP-113,818 significantly inhibited cholesterol absorption, prevented the increase in liver cholesterol, and improved the lipoprotein profiles by selectively decreasing the cholesterol concentrations of the atherogenic lipoproteins (VLDL, IDL, and LDL) in a variety of cholesterol-fed animals. These data suggest that ACAT inhibition may be a useful therapeutic approach for lowering LDL cholesterol and thereby reducing the risk of developing coronary heart disease.