Demonstration of an inwardly rectifying K+ current component modulated by thyrotropin-releasing hormone and caffeine in GH3 rat anterior pituitary cells

 Reduction of an inwardly rectifying K⁺ current by thyrotropin-releasing hormone (TRH) and caffeine has been considered to be an important determinant of electrical activity increases in GH₃ rat anterior pituitary cells. However, the existence of an inwardly rectifying K⁺ current component was recently regarded as a misidentification of an M-like outward current, proposed to be the TRH target in pituitary cells, including GH₃ cells. In this report, an inwardly rectifying component of K⁺ current is indeed demonstrated in perforated-patch voltage-clamped GH₃ cells. The degree of rectification varied from cell to cell, but both TRH and caffeine specifically blocked a fraction of current with strong rectification in the hyperpolarizing direction. Use of ramp pulses to continuously modify the membrane potential demonstrated a prominent blockade even in cells with no current reduction at voltages at which M-currents are active. Depolarization steps to positive voltages at the maximum of the inward current induced a caffeine-sensitive instantaneous outward current followed by a single exponential decay. The magnitude of this current was modified in a biphasic way according to the duration of the previous hyperpolarization step. The kinetic characteristics of the current are compatible with the possibility that removal from inactivation of a fast-inactivating delayed rectifier causes the hyperpolarization-induced current. Furthermore, the inwardly rectifying current was blocked by astemizole, a potent and selective inhibitor of human ether-á-go-go -related gene (HERG) K⁺ channels. Along with other pharmacological and kinetic evidence, this indicates that the secretagogue-regulated current is probably mediated by a HERG-like K⁺ channel. Addition of astemizole to current-clamped cells induced clear increases in the frequency of action potential production. Thus, an inwardly-rectifying K⁺ current and not an M-like outward current seems to be involved in TRH and caffeine modulation of electrical activity in GH₃ cells. Received: 15 May 1997 / Received after revision and accepted: 24 July 1997     

Lysosomal response of a murine macrophage-like cell line persistently infected with Coxiella burnetii.

The lysosomal response of a murine macrophage-like tumor cell line (J774) during persistent infection with Coxiella burnettii was examined. By using acid phosphatase as a lysosomal marker, it was shown that phagosome-lysosome fusion occurred in J774 cells persistently infected with C. burnetii. This observation was verified using thorium dioxide, an electron-dense compound that is sequestered in secondary lysosomes. The phagolysosomes contained viable replicating rickettsiae. Spectrofluorometric analysis indicated that the phagolysosomal pH of persistently infected cells was acidic. In attempts to correlate rickettsial survival with lysosome function, the activities of several lysosomal enzymes were assayed in both infected and uninfected cells. Activities of acid phosphatase and beta-acetylglucosaminidase were not significantly altered during infection. However, infected cells appeared to display slightly higher intracellular lysozyme, beta-glucuronidase, and beta-galactosidase activities.     

Isolation and identification of pathogenic Acanthamoeba strains in Tenerife, Canary Islands, Spain from water sources

A comprehensive survey to document the presence of free-living amoebae of the genus Acanthamoeba was conducted in tap water and sea water sources related to human environments in Tenerife, Canary Islands, Spain. Acanthamoeba identification was based on the morphology of cyst and trophozoite forms and PCR amplification with a genus-specific primer pair. The pathogenic potential of Acanthamoeba isolates was characterized by temperature and osmotolerance assays and PCR reactions with two primer pairs related to Acanthamoeba pathogenesis. The results demonstrate the presence of potentially pathogenic strains in both sources. Thus, some of the amoebae in these aquatic habitats can act as opportunistic pathogens, could play a role in the diseases of aquatic organisms, and may present a risk to human health.     

Familial pericentric inversion of chromosome 8

Eight children from seven presumably unrelated families were identified independently as having an unbalanced recombinant chromosome resulting in the presence of extra material on the short arm of a chromosome 8. Parental chromosomes were analyzed, and one member of each couple (four fathers and three mothers) was found to carry a pericentric inversion of a chromosome 8 [inv(8)(p23q22)]. The propositi had an unbalanced recombinant chromosome [rec(8),dup q,inv(8)(p23q22)]. The affected infants all had developmental delay, congenital heart disease, and unusual appearance. A common origin of the pericentric inversion was suggested because of geographic location and Mexican--American ancestry of the seven families.     

GLP-1 signalling and effects on glucose metabolism in myocytes from type 2 diabetic patients

Changes in the activity of glycogen synthase a and related kinases (phosphatidylinositol-3-kinase, protein kinase B, p44/42 MAP kinases and p70s6 kinase) evoked by GLP-1 in human myocytes from normal subjects were recently implied in the effect of this hormone upon D-glucose transport and glycogen synthesis in the same cells. The major aims of the present study were i) to investigate the possible extension of this knowledge to myocytes obtained from type 2 diabetic patients, ii) to compare in these patients the response to GLP-1, insulin or the structurally related GLP-1 peptides, exendin (1-39)amide and exendin(9-39)amide, and iii) to explore possible differences in the responsiveness to these agents between normal and diabetic subjects. Apart from the much higher basal PI3K activity and impaired response to insulin of p44/42 MAP kinases in the diabetic patients, the changes in enzyme activity caused by either hormone or peptide, although not identical, were essentially comparable. Nevertheless, significant differences in glucose transport and metabolism parameters were observed in the diabetic patients vs. normal subjects: in the diabetic patients, basal 2-deoxy-glucose uptake and glycogen synthase a activity were lower, accompanied by a similar increasing effect of GLP-1 or insulin; yet, the basal value for glycogen synthesis was higher, coinciding with a lesser relative increment in response to GLP-1 or insulin.     

Viral etiopathogenesis of Sjögren's syndrome: role of the hepatitis C virus

Patients with hepatitis C virus (HCV) chronic infection present some extrahepatic manifestations that may mimic the clinical, immunologic and histological manifestations of primary Sj?gren's syndrome (SS). Thus, HCV patients with sicca symptomatology and positive autoantibodies could be misdiagnosed as a 'primary' SS. Nevertheless, there are several clinical and immunologic features that could help us differentiate both processes.     

Correlation between transmission and structure in avian ciliary ganglion synapses

1. Extracellular responses from post-ganglionic axons of pigeon and chick isolated ciliary ganglia were elicited by stimulation of the presynaptic nerve. Intracellular recordings were also obtained from newly hatched pigeon and chick ganglion cells. The fine structure of ganglia from pigeons of various ages was examined with the electron microscope.2. In ganglia from chick embryos and pigeons up to 10 days old, the extracellular response was unimodal with a long latency and could be blocked by the addition of D-tubocurarine (D-TC) or hexamethonium to the bathing solution. A bimodal extracellular response appeared in pigeons about 10 days after hatching. Only the second peak of the response could be blocked by D-TC or hexamethonium. The response recorded from 22 to 26-day-old pigeons was similar to that seen in the adult.3. The intracellular recordings from ganglion cells of 2-week-old pigeons exhibit two post-synaptic potentials elicited by presynaptic stimulation. The first post-synaptic potential appears to be due to current flow through the ganglion cell during the presynaptic action potential. The second is chemically mediated. In pigeons from 1 to 6 days old, only the second post-synaptic potential is observed.4. The presynaptic terminals in the 4-day-old birds were in the form of calyces. In pigeons 7 days old or older, boutons appeared. The boutons were presumably formed as a result of cleavage of calyciform nerve terminals. Myelin was seen first in the 7-day-old pigeon, was well developed in the 16-day-old bird, and persisted in the adults.5. In adult ganglia, the first component of the extracellular response decreased and was finally abolished after 10-12 hr of superfusion with Tyrode solution. The second component of the response increased concomitantly. The only anatomical change noted in the ganglia after soaking was the disruption and separation of the myelin lamellae from each other and from around the ganglion and presynaptic terminals.6. It is concluded that the myelin is necessary for electrical transmission in the pigeon ciliary ganglion.