Glucagon-like peptide 1 receptor expression in primary porcine proximal tubular cells.

BACKGROUND: GLP-1 is secreted into the circulation after food intake. The main biological effects of GLP-1 include stimulation of glucose dependent insulin secretion and induction of satiety feelings. Recently, it was demonstrated in rats and humans that GLP-1 can stimulate renal excretion of sodium. Based on these data, the existence of a renal GLP-1 receptor (GLP-1R) was postulated. However, the exact localization of the GLP-1R and the mechanism of this GLP-1 action have not yet been investigated. METHODS: Primary porcine proximal tubular cells were isolated from porcine kidneys. Expression of GLP-1R was measured at the mRNA level by quantitative RT-PCR. Protein expression of GLP-1R was verified with immunocytochemistry, immunohistochemistry and Western blot analysis. Functional studies included transport assessments of sodium and glucose using three different GLP-1 concentrations (200 pM, 2 nM and 20 nM), 200 pM exendin-4 (GLP-1 analogue) and an inhibitor of the dipeptidylpeptidase IV (DPPIV) enzyme (P32/98 at 10 microM). Finally, the expression of NHE3, the predominant Na(+)/H(+) exchanger in proximal tubular cells, was also investigated. RESULTS: GLP-1R, NHE3 and DPPIV were expressed at the mRNA level in porcine proximal tubular kidney cells. GLP-1R expression was confirmed at the protein level. Staining of human and pig kidney cortex revealed that GLP-1R was predominantly expressed in proximal tubular cells. Functional assays demonstrated an inhibition of sodium re-absorption with GLP-1 after 3 h of incubation. Exendin-4 and GLP-1 in combination with P32/98 co-administration had no clear influence on glucose and sodium uptake and transport. CONCLUSION: GLP-1R is functionally expressed in porcine proximal tubular kidney cells. Addition of GLP-1 to these cells resulted in a reduced sodium re-absorption. GLP-1 had no effect on glucose re-absorption. We conclude that GLP-1 modulates sodium homeostasis in the kidney most likely through a direct action via its GLP-1R in proximal tubular cells.

Efficacy of crisis intervention.

This study assesses the efficacy of the treatment approach implemented in the Bern Crisis Intervention Program, where particular emphasis is placed on the remediation of suicide ideation and suicidal behavior, and depression, fear, and phobia are generally considered to be contributing factors. Four questionnaires addressing psychopathology, emotional well-being, social anxiety, and personality were administered prior to and after the treatment of 51 patients over a period of 2 to 3 weeks. The reduction of symptoms contributing to suicidal ideation and behavior was interpreted as indirect evidence of an antisuicidal effect of the program. Significant improvements were found in the psychopathology ratings, with depression and anxiety showing the largest reductions. The impact on personality and social phobia, however, was only moderate, and on average patients still exhibited symptoms after attending the program. This residual symptomatology points to the necessity of introducing a two-step therapy approach of intensive intervention targeted at the precipitating causes of the crisis, augmented by long-term therapy to treat underlying problems.

Quantitative study of intussusceptive capillary growth in the chorioallantoic membrane (CAM) of the chicken embryo.

In an attempt to sort out the respective contributions of sprouting and intussusceptive microvascular growth (IMG) during chicken chorioallantoic membrane (CAM) development, we analyzed the morphology and the quantitative growth of the capillary bed of the CAM by light microscopy. By perfusing the CAM microvasculature with highly concentrated colloidal gold particles, the capillaries could be unambiguously distinguished from the surrounding unlabelled tissue. This allowed us to identify, count and measure the intercapillary tissue profiles. By means of morphometric analysis we could show that CAM angiogenesis undergoes three phases of development. In an early phase, from Day 5 to Day 7, the major mechanism of capillary network growth is sprouting. In an intermediate phase, from Day 8 to Day 12, IMG is prevailing, and at Days 13 and 14, CAM structure is undergoing expansion with only a small increase in complexity. These findings are important in view of experimental protocols using the CAM as a model for testing angiogenetic factors. Indeed, care has to be taken not to misinterpret normal age-dependent alterations of the CAM vascular architecture as specific responses to tested agents.

Characterization of a Ca(2+)-dependent anion channel from sheep tracheal epithelium incorporated into planar bilayers.

1. Anion-selective channels from the apical membrane of respiratory epithelia are involved in the secretion of chloride into the airway lumen. In cystic fibrosis (CF) there is an abnormality of phosphorylation-regulated chloride transport in this tissue, whilst a calcium-dependent pathway appears to function normally. 2. Using incorporation of apical membrane vesicles into planar phospholipid bilayers, we have characterized the most commonly seen anion-selective channel from sheep tracheal epithelium. 3. In symmetrical 200 mM-NaCl solutions the channel showed rectification, with a chord conductance at negative voltages of 107 pS and at positive voltages of 67 pS. The channel characteristically demonstrated subconductance states at 1/3 and 3/4 of the fully open level. Selectivity for chloride over sodium was approximately 6:1. 4. The channel required a minimum of approximately 100 microM-calcium on the presumed cytoplasmic surface (cis) for opening events to be observed. Open probability (Po) of the fully open state was markedly voltage dependent, but little effect of voltage was seen on the 1/3 subconductance state. 5. The relative permeabilities of monovalent anions monitored under bi-ionic conditions gave the following sequence: NO3- greater than I- greater than Cl- = Br- much much greater than F-. The order of conductances in symmetrical solutions was Cl- = NO3- greater than Br- greater than I- much much greater than F-. 6. The chloride channel blocker 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) produced a dose-related reduction in Po with a flickering block at 10-50 microM and complete block at higher concentrations. 7. ATP produced a dose-related reduction in Po with effects at 1 microM and complete closing at 1 mM. These effects were only seen with addition to the cis chamber. 8. The catalytic subunit of protein kinase A, either when incubated with vesicles prior to incorporation into bilayers, or when added directly to either chamber, produced no effect. 9. Channels with very similar properties were seen from transfected human tracheo-bronchial cells. 10. Recent whole-cell patch-clamp studies have suggested a distinct calcium-activated chloride current in secretory epithelia. The described channel has properties in common with this current and may be a candidate for its single-channel basis.