Activation of the tubuloglomerular feedback mechanism in dehydrated rats

To study the influence of the tubuloglomerular feedback control (TGF) on the regulation of glomerular filtration rate (GFR) during dehydration, micropuncture experiments were performed on surface nephrons of dehydrated rats. Dehydration was achieved by withdrawal of food and water for 24 h. The urine flow rate decreased to 1.5 μl/min (controls 2.9 μl/min) and GFR decreased in these rats to 0.80 ml/min (controls 1.22). TGF was studied by two different micropuncture procedures. With the first technique the changes in proximal stop-flow pressure in response to changes of the late proximal microperfusion rate were measured. With this technique the perfusion rate necessary to induce a half maximal stop-flow pressure response, the turning point, was also determined. An increased TGF sensitivity was found in dehydrated rats, as indicated by increased stop-flow pressure responses (35 versus 26%) and decreased turning points (16 versus 21 nl/min). With the second micropuncture technique the single nephron GFR (SNGFR) was measured at distal and proximal tubular sites, in the same nephron. Distal SNGFR was decreased during dehydration to 32.2 nl/min, versus 42.7 nl/min in controls. A significant difference between paired SNGFR measurements in the same nephron was observed during dehydration, the proximal value being 5.3 nl/min higher than the distal, whereas this difference was not seen in control rats. This finding indicates that activation of the feedback mechanism takes place to reduce SNGFR. It is concluded that the decrease in whole kidney GFR is partly caused by the observed increase in feedback activity. The present results are also in agreement with our earlier hypothesis that the hydrostatic and oncotic pressure conditions within the interstitial space surrounding the macula densa cells modulate the sensitivity of the tubuloglomerular feedback mechanism.     

Susceptibility of the sympathectomized ear to noise-induced hearing loss

Unilaterally sympathectomiced rats were exposed to 100 dB L_eq frequency-modulated noise for I month. Normotensive as well as spontaneously hypertensive animals (with a blood pressure of above 200 mmHg) were investigated. Auditory sensitivity was determined by auditory brainstem responses to 1/3-octave filtered sine waves in the frequency range 0.8–20.0 kHz. In addition, a morphological analysis was carried out. It was found that the sympathetic innervation to the inner ear of the rat originated in, or passed through the ipsilateral superior cervical ganglion. Sympathectomy did not alter pre-exposure hearing thresholds nor influence the size of the noise-induced hearing loss either in 3 or 11 months old hypertensive rats, or in normotensive rats of 11 months. A slightly smaller loss was seen in the sympathectomized side in young normotensive rats. It was concluded that the sympathetic does not exert a protection of the inner ear against functional disturbances in hypertension, neither during basal metabolic condition nor during extreme conditions, i.e. during noxious noise exposure.     

Brain extracellular space during spreading depression and ischemia

The change of extracellular space volume of rat brain cortex during ischemia and cortical spreading depression, CSD (Leāo4) was evaluated by a new method. The cortical surface was imgated with isotonic CSF containing the extracellular markers 50 mM cholin or 50 mM trimethyltris(hydroxymethyl)methyl ammonium ion (N-TRIS), and their extracellular concentrations were monitored by ion-selective microelectrodes. When steady-state for the concentration of these markers was attained, CSD evoked a reversible increase of the concentration of the markers, indicating shrinkage of the interstitial volume of distribution. During ischemia an initial slow rate of concentration increase was observed, followed a few minutes later by a rapid increase concomitant with the sharp rise in extracellular potassium concentration. During CSD and ischemia, the maximal increases of choline and N-TRIS concentration reflected a shrinkage of the extracellular space amounting to about 50% of the initial volume.     

Nuclear magnetic resonance studies of the C-terminal human growth hormone fragment I179–C182–[SS]-C189-P191 and the related trisulfide peptide I179–C182-[SSS]-C189–P

The synthetic C-terminal hGH fragment I179-C182-[SS]-C189-P191 and the related trisulfide peptide I179-C182-[SSS]-C189-P191 have been studied using homonuclear ¹H-NMR methods and distance geometry calculations. The ¹H-NMR spectra of both the disulfide (diS) and the trisulfide (triS) were completely assigned. Amide proton exchange rates, NOEs and the temperature dependence of the NH chemical shifts indicate a hydrogen bond in triS between Val 185 and Ser 188 stabilizing a turn in this region. ³J_{H, H} coupling constants and NOEs were measured and used as input for distance geometry calculations. For triS two families of structures with averaged pairwise backbone root mean square deviations for Cys 182-Cys 189 of 1.3–1.5 Å were found, only one of which is compatible with experimental data. For diS only one family of structures was found, but with such a low structural definition (backbone rmsd >2Å) that no interpretation into a consensus structure is useful. The generated structures were compared to the crystal structure of the terminal loop in hGH. complexed to its binding proteins. The resemblance was low between the solution structures of the tridecapeptides and the terminal hGH loop. © Munksgaard 1997.     

Unexpected lability of cysteine acetamidomethyl thiol protecting group Tyrosine ring alkylation and disulfide bond formation upon acidolysis

Cleavage and deprotection of the peptidyl resin H-Asn-Gly-Gly-Cys(Acm)-Glu(OBu^t)-Gln-Tyr(Bu^t)-Cys(Acm)-Ser(Bu^t)-Asp(OBu^t)-[(p-alkoxy)benzyloxy polystyrene resin] using standard conditions with various trifluoroacetic acid-containing mixtures were found to result in partial removal of ordinarily acid-stable S-Acm groups. Thus, apart from the desired peptide H-Asn-Gly-Gly-Cys(Acm)-Glu-Gln-Tyr-Cys(Acm)-Ser-Asp-OH, a disulfide-cyclic peptide derivative was also isolated. Furthermore, it was found that in another major by-product of the peptide resin cleavage the tyrosine side chain had been alkylated with an Acm group in a position ortho to the phenolic function. The formation of both by-products could be suppressed by carrying out the cleavage/deprotection reaction at higher dilution and by inclusion of scavengers such as phenol. An authentic sample of the disulfide-cyclic peptide was obtained by oxidation of H-Asn-Gly-Gly-Cys-Glu-Gln-Tyr-Cys-Ser-Asp-OH using Ellman's reagent. © Munksgaard 1997.