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K S Cuthbertson

Publications and source records attributed to K S Cuthbertson.

29 records · Page 2Linked to original sources
PubMed1987

Phorbol-ester-induced alterations of free calcium ion transients in single rat hepatocytes.

The effect of the phorbol esters phorbol 12-myristate 13-acetate (TPA) and phorbol 12,13-dibutyrate (PDB) on changes in free Ca2+ concentration ([Ca2+]i) in single rat hepatocytes, microinjected with the photoprotein aequorin, were investigated. [Arg8]vasopressin and phenylephrine induced a series of repetitive [Ca2+]i transients. Phorbol esters inhibited the alpha 1-adrenoceptor-induced response; sub-nanomolar concentrations decreased the transient frequency, and higher concentrations abolished the transients. The inhibitory effect of PDB was readily reversible. Phorbol esters were less effective in decreasing the frequency of [Arg8]-vasopressin-induced transients, and the inhibition could be overcome by high [Arg8]vasopressin concentrations.

Aequorin↗
PubMed1987

Cytosolic free Ca2+ in single rat heart cells during anoxia and reoxygenation.

Free Ca2+ in the cytosol ([Ca2+]i) of individual rat ventricle cells injected with aequorin was measured under anoxia. In glucose-free medium myocytes spontaneously shortened after about 60 min, although [Ca2+]i was still at or near resting levels. However, within minutes a net inward movement of Ca2+ across the sarcolemma developed and [Ca2+]i began to rise. Provided oxygen was readmitted before [Ca2+]i exceeded 2-3 microM, cells were able to restore [Ca2+]i to resting levels through caffeine-sensitive sequestration of Ca2+ in the sarcoplasmic reticulum. We suggest that Ca2+-independent shortening of anoxic cardiomyocytes reflects onset of rigor which triggers loss of [Ca2+]i homoeostasis.

Animals↗
PubMed1987

Calcium transients in single adrenal chromaffin cells detected with aequorin.

The effect of 55 mM K+ and nicotine on intracellular free calcium was monitored in bovine adrenal chromaffin cells microinjected with aequorin. In contrast to results with quin 2, which suggested that stimulation of chromaffin cells resulted in sustained rises in free calcium, aequorin measurements showed that 55 mM K+ and nicotine resulted in a transient (60-90 s) elevation of free calcium. The peak free calcium and duration of the transient elicited by nicotine were dose-dependent. The concentration of nicotine (10 microM) giving a maximal secretory response gave a peak rise in free calcium of up to 1 microM. 55 mM K+ which only releases 30% of the catecholamine released by 10 microM nicotine generated a calcium transient indistinguishable from that due to 10 microM nicotine. These results support the idea that nicotine agonists generate an alternative second messenger in addition to the rise in free calcium.

Adrenal Medulla↗
PubMed1987

Agonist-induced oscillations in cytoplasmic free calcium concentration in single rat hepatocytes.

The effects of the alpha 1-adrenergic agonist phenylephrine and the peptide hormones angiotensin II and arg8-vasopressin on cytoplasmic free calcium concentration were investigated in single rat hepatocytes microinjected with the photoprotein aequorin. Hepatocytes responded to physiological concentrations of the glycogenolytic agonists with a series of repetitive Ca transients. In each transient free Ca rose in 2-3s to above 600 nM from a resting level of 200 nM. Transient duration depended on the agonist and ranged from approximately 7s for phenylephrine to approximately 15s for angiotensin. Transient frequency, but not shape or size, depended on agonist concentration. The period ranged from less than 20s to several minutes. We suggest that the frequency of the Ca transients is the principal determinant of the amplitude of the cellular response to calcium-mobilizing agonists.

Aequorin↗
PubMed1983

Aequorin measurements of free calcium in single mammalian cells.

The calcium-triggered photoprotein aequorin has been used as a sensitive indicator of cytoplasmic free calcium in cells of relatively large size. We describe here several novel modifications to the technique that make it possible to detect resting levels of free calcium in single mammalian oocytes, fibroblasts and granulosa cells. Concentrations of free calcium as low as 1 X 10(-7) M have been measured in single fibroblasts, corresponding to a detection limit of 10(-18)mol.

Aequorin↗
PubMedDate not supplied

Modelling receptor-controlled intracellular calcium oscillators.

This paper presents mathematical models for the hepatocyte calcium oscillator which follow the concepts in a class of informal models developed to account for the striking dependence on the receptor type of several features of the calcium oscillations, in particular the shape and duration of the free calcium transients. The essence of these models is that the transients should be timed by a build-up of activated GTP-binding proteins, which, combined with positive feedback processes and perhaps with cooperative effects, leads to a sudden activation of phospholipase C (PLC), followed by negative feedback processes which switch off the calcium rise and lead to a fall in free calcium back to resting levels. These models predict pulsatile oscillations in inositol (1,4,5)P3 as well as in free calcium. We show that receptor-controlled intracellular calcium oscillators involving an unknown positive feedback pathway onto PLC and negative feedback from protein kinase C (PKC) onto G-proteins and receptors, or negative feedback by stimulation of GTPase activity can simulate many of the features of observed intracellular calcium oscillations. These oscillators exhibit a dependence of frequency on agonist concentration and a dependence of transient duration on receptor and G-protein type. We also show that a PLC-dependent GTPase activating factor (GAF) could provide explanations for some otherwise puzzling features of intracellular calcium oscillations.

Animals↗
PubMedDate not supplied

Phorbol ester and sperm activate mouse oocytes by inducing sustained oscillations in cell Ca2+.

The parthenogenetic activation of mouse oocytes by several agents is accompanied by a large rise in the intracellular calcium concentration ( [Ca2+]i). The tumour-promoting phorbol ester 12-O-tetradecanoyl phorbol acetate (TPA) activates some cellular processes which are also activated by raised [Ca2+] (ref. 2). We therefore tested TPA on mouse oocytes and found it to be a potent parthenogenetic activating agent. Recent reports suggest that TPA mimics endogenous diacylglycerol3 and can stimulate cells by activating protein kinase C without raising [Ca2+]i (refs 4, 5). We have now measured [Ca2+]i with aequorin in mouse oocytes treated with TPA. Sustained oscillations in [Ca2+]i appeared in those oocytes which were subsequently activated. We also measured [Ca2+]i during fertilization. The response began with [Ca2+]i transients as reported previously, but the transients continued in a regular pattern for 4 h. There was no rise in [Ca2+]i of the type induced by activating agents such as ethanol. One component of the fertilization response appears to be related to the TPA-induced oscillations.

Animals↗
PubMedDate not supplied

Repetitive transient rises in cytoplasmic free calcium in hormone-stimulated hepatocytes.

In the stressed animal, the vasoactive hormones vasopressin and angiotensin-II and the neurotransmitter noradrenaline induce liver cells to release glucose from glycogen. The intracellular signal that links the cell-surface receptors for noradrenaline (alpha 1) and vasoactive peptides to activation of glycogenolysis is known to be a rise in the cytoplasmic concentration of free calcium ions (free Ca). The receptors for these agonists induce the hydrolysis of phosphatidylinositol 4,5-bisphosphate, a minor plasmalemma lipid, to produce inositol trisphosphate and diacylglycerol. Inositol trisphosphate has been shown to mobilize intracellular calcium in hepatocytes. We show here, by means of aequorin measurements in single, isolated rat hepatocytes, that the free Ca response to these agonists consists of a series of transients. Each transient rose within 3 s to a peak free Ca of at least 600 nM and had a duration of approximately 7 s. The transients were repeated at intervals of 0.3-4 min, depending on agonist concentration. Between transients, free Ca returned to the resting level of approximately 200 nM. Clearly, the mechanisms controlling free Ca in hepatocytes are more complex than hitherto suspected.

Aequorin↗