Human Brain‐Type Glycogen Phosphorylase: Quantitative Localization in Human Tissues Determined with an Immunoassay System

Abstract

Glycogen phosphorylase (EC 2.4.1.1) from human brain tissue was purified to homogeneity. Antisera were developed in rabbits with purified phosphorylase as the immunogen. Antibodies were first affinity-purified with a column of brain phosphorylase-coupled Sepharose, and then the antibody fraction was adsorbed with a column of muscle phosphorylase-coupled Sepharose to remove antibodies reactive also with muscle phosphorylase. By using the specific antibodies, a sandwich-type immunoassay system for measurement of brain phosphorylase was prepared. The assay system consisted of polystyrene balls with immobilized antibrain phosphorylase F(ab')2 fragments and the same antibody Fab' fragments labeled with beta-D-galactosidase from Escherichia coli. The assay was sensitive and specific to the brain phosphorylase. The minimum detection limit of the assay was 0.1 ng/assay tube, and the cross-reactivity of the assay with muscle phosphorylase was less than 1%. Tissue concentrations of immunoreactive brain-type phosphorylase were estimated. The phosphorylase was present in the heart at as high a level as in the brain. The immunoreactivity for brain phosphorylase was distributed widely at a significant concentration in various peripheral tissues, such as the digestive tract, bladder, aorta, liver, and testis. Immunohistochemical localization of brain phosphorylase in the CNS revealed that the enzyme is present in most astrocytes and amyloid bodies, as well as in some neurons in the cerebral cortex and Golgi cells in the cerebellar cortex.

Mode of expression of brain- and muscle-type glycogen phosphorylase in extraocular muscles of human embryos.

“…The preparation procedures and the specificity of the antibodies used in the present study are described in another paper (5). BGP antibody is specific to brain-type phosphorylase and MB-GP antibody is common to brain and muscle phosphorylase (5). BGP and MB-GP showed no activity in liver cells in all embryos examined, suggesting that these antibodies do not crossreact with LGP.…”

“…Serial sections of 5 p-thickness were prepared and were immunostained after avidin-biotin- (4), using BGP and MB-GP antibodies. The preparation procedures and the specificity of the antibodies used in the present study are described in another paper (5). BGP antibody is specific to brain-type phosphorylase and MB-GP antibody is common to brain and muscle phosphorylase (5).…”

A Preferential Role for Glycolysis in Preventing the Anoxic Depolarization of Rat Hippocampal Area CA1 Pyramidal Cells

“…2C,D). If we ignore metabolic compartmentation within the slice by assuming that energy stored as glycogen predominantly in glia (Rosenberg and Dichter, 1985;Kato et al, 1989;Ignacio et al, 1990) can be freely passed to neurons (e.g., as lactate) (Cambray-Deakin et al, 1988;Dringen et al, 1993;Pellerin et al, 1998;Sibson et al, 1998;Magistretti and Pellerin, 1999) and also assume that with mitochondria blocked there is no change in the rate of ATP consumption or in the time needed for ion gradients to decrease after the sodium pump has stopped, Figure 9. Response of CA1 pyramidal cells to oxygen and glucose deprivation without metabolic blockers.…”

Stimulation of energy metabolism by α‐adrenergic agonists in primary cultures of astrocytes

“…A continuously increasing amount of evidence suggests that astrocytes also in vivo have a high density of adrenergic receptors (reviewed in Hertz, 1989b;Stone and Ariano, 1989). A confinement of the metabolic stimulation to astrocytes is in conceptual agreement with the findings that glycogen, glycogen phosphorylase activity, and noradrenaline-stimulated glycogenolysis are localized in astrocytes (Ibrahim, 1975;Cummins et al, 1983a,b;Cambray-Deakin et al, 1988;Kala et al, 1989;Kato et al, 1989). If astrocytes are, indeed, the major target, for noradrenergic stimulation of brain metabolism, the stimulation can, however, be expected secondarily to affect also neurons, and thus brain output, on account of pronounced metabolic and functional interactions (Hertz, 1989a,b) between astrocytes and neurons.…”