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Biomedical subjects

T Maack

Publications and source records attributed to T Maack.

At least 37 records · Page 2Linked to original sources

Endocytosis and lysosomal hydrolysis of proteins in proximal tubules.

The techniques and basic protocols described above can be readily reproduced by investigators with experience in perfusion of isolated nephron segments. They can be modified and adapted by the investigator to address specific issues. In particular, isolated perfused nephron segments have also been successfully used for elucidation of biochemical and morphological aspects of endocytosis and lysosomal hydrolysis of macromolecules, proteins, and polypeptides. The reader is directed to the references cited under Methodological Approaches in this chapter for a description of these techniques. Although studies on the endocytic uptake and metabolism of proteins and polypeptides using isolated perfused nephron segments have made significant inroads in our understanding of these fascinating and important biological processes, much remains to be learned. Hopefully, future uses of the technique will further advance our knowledge in this field.

Absorption↗

Atrial natriuretic factor receptors in cultured renomedullary interstitial cells.

To gain further insight on the cell types that may mediate the effects of atrial natriuretic factor (ANF) in the renal medulla, we determined the distribution and function of biological (B) and clearance (C) receptors of ANF in renomedullary interstitial cells (RMIC). Studies were performed in the 3rd-17th passages of RMIC obtained from a primary culture of the rat renal medulla. Electron microscopy of the cultured cells showed the typical morphological features of RMIC "in vivo," including prominent lipid droplets. RMIC have a very high density of high-affinity specific binding sites of ANF-(1-28) [23,000 sites/cell; dissociation constant (Kd) = 50 pM]. There was only minimal binding of C-ANF-(4-23) (less than 2,500 sites/cell), a specific ligand of C-ANF receptors. ANF-(1-28) markedly increased guanosine 3',5'-cyclic monophosphate (cGMP) from 1.3 +/- 0.3 to 106 +/- 22 pmol cGMP/10(6) cells [50% effective dosage (ED50) = 1.2 nM]. The effect of ANF-(1-28) on cGMP was nearly additive to that of sodium nitroprusside and was not potentiated or antagonized by C-ANF-(4-23). The density of guanylate cyclase-coupled B-ANF receptors and the ANF-induced increase in cGMP in RMIC are higher than those reported to date in other target cells. This suggests that RMIC may mediate some of the known effects of ANF in the renal medulla.

Animals↗

Clearance function of type C receptors of atrial natriuretic factor in rats.

The overwhelming majority of atrial natriuretic factor (ANF) receptors in kidney and vascular tissues do not mediate any of the known functional effects of the hormone. To test whether these receptors (C-ANF receptors) function as clearance receptors for circulating ANF-(1-28), we determined the effects of C-ANF-(4-23) [des[Gln18Ser19Gly20Leu21Gly22]rANF-(3-23)-NH2], a specific ligand of C-ANF receptors, on the pharmacokinetics and hydrolysis of 125I-labeled ANF-(1-28) in anesthetized rats. Radioactivity in plasma was characterized by trichloroacetic acid solubility and high-pressure liquid chromatography. C-ANF-(4-23) (1 and 10 micrograms.min-1.kg body wt-1) led to marked dose-dependent increases in initial plasma concentration of administered 125I-ANF-(1-28) and decreases in its volume of distribution at steady state (Vss), metabolic clearance rate (MCR), and appearance of hydrolytic products ([125I]monoiodotyrosine and free 125I) in plasma (Pm). At the highest dose, C-ANF-(4-23) decreased Vss from 97 +/- 12 to 36 +/- 2 ml/100 g body wt, MCR from 50 +/- 4 to 12 +/- 1 ml.min-1.100 g body wt-1, and Pm from 54 +/- 8 to 11 +/- 2% of initial plasma 125I-ANF-(1-28). The data demonstrate that C-ANF receptors are mainly responsible for the very large volume of distribution and fast MCR of ANF in the rat. In this manner, C-ANF receptors are likely to play an important role in the homeostasis of circulating ANF.

Animals↗

Physiological role of silent receptors of atrial natriuretic factor.

A ring-deleted analog of atrial natriuretic factor--des[Gln18, Ser19, Gly20, Leu21, Gly22] ANF4-23-NH2 (C-ANF4-23)--binds with high affinity to approximately 99% of ANF receptors in the isolated perfused rat kidney. In this preparation, C-ANF4-23 is devoid of detectable renal effects and does not antagonize any of the known renal hemodynamic and natriuretic actions of biologically active ANF1-28. In contrast, both C-ANF4-23 and ANF1-28 increase sodium excretion and decrease blood pressure in intact anesthetized rats. This apparent contradiction is resolved by the finding that the ring-deleted analog markedly increases plasma levels of endogenous immunoreactive ANF in the rat. The results show that the majority of the renal receptors of ANF are biologically silent. This new class of receptors may serve as specific peripheral storage-clearance binding sites, acting as a hormonal buffer system to modulate plasma levels of ANF.

Animals↗

Binding and functional effects of atrial natriuretic factor in isolated rat kidney.

A new methodological approach was developed to study the relationship between specific binding and dose-response curves of the renal effects of atrial natriuretic factor (ANF) in isolated perfused rat kidneys (IK). IK were perfused with 125I-labeled and unlabeled ANF 1-28 (4 pM to 1 microM) to determine the following: 1) distribution, capacity (Cmax), and apparent affinity (S50) of specific binding of ANF 1-28 in cortex, outer medulla, and papilla and 2) dose-response curves of the effects of ANF 1-28 on renal hemodynamics and excretion of fluid and electrolytes. The kidney had a very high density of high-affinity binding sites for ANF. Cortex had greater than 90% of total binding sites (Cmax = 6.8 pmol/g tissue; S50 = 54 pM), whereas papilla had less than 2% of total binding sites with a 10-fold lower apparent affinity (S50 = 860 pM) than in cortex. ANF-induced increases in glomerular filtration rate and excretion of fluid and electrolytes were detectable at 10-100 pM and maximal effects occurred at 1-10 nM ANF. Below 1 nM there was no dissociation between the renal hemodynamic and natriuretic effects of ANF. There was a close agreement between dose-response and binding curves of ANF to cortex. Results demonstrate that binding site occupancy in kidney cortex and renal effects of ANF occur at near physiological concentrations of the hormone.

Animals↗

Effects of atrial natriuretic factor on the kidney and the renin-angiotensin-aldosterone system.

ANF is a peptide hormone with peripheral and central effects on several physiologic control systems, which suggests broad involvement in the regulation of intravascular volume and cardiovascular homeostasis. ANF acts directly on the kidney to modulate renal vascular resistance, increase glomerular filtration rate, and decrease inner medullary hypertonicity. These hemodynamic effects act in concert to promote marked natriuresis and diuresis and to inhibit renin secretion by the kidney; the potential role of direct effects on tubular transport and on the juxtaglomerular cells remains to be clarified. ANF also inhibits steroidogenesis, most prominently affecting agonist-induced aldosterone biosynthesis by the adrenal cortex. ANF exerts a smooth muscle relaxant effect on isolated vessels constricted with various hormonal agonists. ANF causes especially pronounced antagonism of the adrenal and vascular actions of angiotensin II and antagonizes the latter peptide's central nervous system effects. Although complex systemic hemodynamic mechanisms are involved, ANF-induced vasorelaxation may contribute to its depressor action, particularly in states characterized by angiotensin II-induced vasoconstriction. Distention of the atria is a major stimulus to ANF release and evokes many responses that are mimicked by ANF infusion. Although ANF is not the sole or even dominant mediator of the responses to atrial distention, available data suggest that it plays a role in the renal and hormonal responses to intravascular volume expansion. Definitive assessment of its physiologic and pathophysiologic significance must await development of specific antagonists, but further studies on the mechanisms of action of ANF at the molecular, cellular, and systemic levels are likely to contribute significantly to our understanding of the complex process of volume regulation and cardiovascular homeostasis.

Animals↗

Renal vasoconstrictive effect of kinins mediated by B1-kinin receptors.

The nature of the renal vascular actions of kinins, their dependence on prostaglandins and B1-kinin receptor responses were studied in functioning isolated perfused rat kidneys (IK). Lysylbradykinin (LBK), 0.28 and 0.7 microM, transiently decreased and then markedly increased the renal vascular resistance (RVR) in a sustained manner. Bradykinin (BK) at the same doses also had a transient vasorelaxant but not a sustained vasoconstrictive effect. The inactivation of LBK and BK by the IK did not account for the transient nature of their vasorelaxant effect. Indomethacin (5 microM) markedly blunted LBK-induced decrease but not increase in RVR. The B1-kinin receptor agonist desArg9-BK (0.4-1.0 microM) did not decrease RVR but, as LBK, markedly increased RVR in a dose-related manner. The B1-kinin receptor antagonist [Leu8]desArg9-BK had no effect on its own but inhibited the desArg9-BK-induced vasoconstriction in a stoichiometric manner. This antagonist at 4.0 microM also completely abolished the vasoconstrictive effect of 0.7 microM LBK, whereas it potentiated and prolonged its vasorelaxant effect. The results demonstrate that kinins, particularly LBK, have bimodal effects on the renal vascular resistance of the isolated perfused rat kidney. The vasorelaxant effect is at least partly mediated by prostaglandins whereas the vasoconstrictive effect of LBK and/or its renal metabolites has the typical character of a B1-kinin receptor response. It is postulated that B1-kinin receptor responses may be of importance in the generation and/or maintenance of renal vasoconstriction in disease states which lead to renal failure.

Animals↗

Atrial natriuretic factor increases hematocrit and decreases plasma volume in nephrectomized rats.

To test the hypothesis that ANF increases the efflux of fluid from capillaries we determined the effect of synthetic (1-28) ANF on hematocrit (Hct) and plasma volume (PV) in 24 hr bilaterally nephrectomized rats. Results were compared with those obtained in bilaterally nephrectomized rats receiving saline alone (S) or an infusion of sodium nitroprusside (NP) to produce a similar blood pressure lowering effect as ANF. PV was determined by the volume of distribution of RISA 10 minutes after its i.v. administration. After control periods (C) rats were infused for 40 minutes with saline alone (group A, n = 8), (1-28)ANF, 0.3 micrograms X min-1/kg body wt (group B, n = 9), 1.0 microgram X min-1/kg body wt (group C, n = 8) and NP, 1.0 to 2.0 micrograms X min-1/kg body wt (group D, n = 8). Recovery periods (R) were performed 45 minutes after the experimental periods (E). During E periods, mean arterial blood pressure did not change in group A and fell significantly (p less than 0.01) in the other groups. Group B: C = 89 +/- 4; E = 82 +/- 4 mmHg. Group C: C = 106 +/- 7; E = 92 +/- 7 mmHg. Group D: C = 104 +/- 5; E = 90 +/- 5 mmHg. Neither saline or NP changed Hct, whereas, ANF significantly (p less than 0.01) increased this parameter (Group B: C = 43.4 +/- 0.7; E = 46.3 +/- 0.6% and Group C: C = 44.5 +/- 1.2; E = 47.4 +/- 1.2%). ANF significantly decreased PV (3.86 +/- 0.14 ml/100 gm body wt) as compared to S (4.51 +/- 0.22 ml/100 gm body wt; p less than 0.05) or NP (4.69 +/- 0.11 ml/100 gm body wt; p less than 0.01). The average decrease in PV of 14% corresponded to the average increase in Hct of 7%. Results demonstrate that ANF increases the efflux of fluid from capillaries. This effect explains, at least in part, the ANF-induced decrease in cardiac output and blood pressure in normotensive animals and may play an important role in the homeostatic regulation of plasma volume.

Animals↗

Role of increased glomerular filtration rate in atrial natriuretic factor-induced natriuresis in the rat.

One of the major renal hemodynamic actions of atrial natriuretic factor (ANF) is to increase glomerular filtration rate (GFR). To assess the role of this effect on ANF-induced natriuresis (UNaV), diuresis (V) and kaliuresis (UKV) we performed late clamp experiments in six rats. After control periods (C), synthetic ANF (auriculin A) was infused i.v. (2 micrograms X min-1/kg body wt) throughout the experiment (150 min). After pre-clamp periods, the perfusion pressure of the left kidney (LK) was reduced to 75-80 mmHg. The right kidney (RK) served as a time control. In LK, before the late clamp, ANF increased (p less than 0.01) GFR from 1.5 +/- 0.1 to 1.8 +/- 0.1 ml/min, V from 17 +/- 5 to 53 +/- 5 microliters/min, and UNaV from 2.1 +/- 0.6 to 10.0 +/- 0.9 microEq/min. Almost identical increases occurred in the RK. The late clamp returned all parameters in LK to C values (p greater than 0.05): GFR to 1.4 +/- 0.1 ml/min, V to 6.3 +/- 1.2 microliter/min, and UNaV to 1.0 +/- 0.3 microEq/min. The late clamp also reversed the ANF-induced increase in UKV. In the RK, GFR (1.8 +/- 0.1 ml/min), V (38 +/- 4 microliter/min) and UNaV (7.8 +/- 0.8 microEq/min) remained elevated (p less than 0.01 vs. C) to the end of the experiment. These data demonstrate that upon return of GFR to control levels, the ANF-induced diuresis, natriuresis and kaliuresis is abolished. The results support our previous view that the increase in GFR together with a decrease in inner-medullary hypertonicity account wholly or in great part for the natriuretic action of ANF.

Animals↗

Expression and secretion of biologically active human atrial natriuretic peptide in Saccharomyces cerevisiae.

A hybrid gene was constructed containing a fusion between the DNA sequences encoding the secretory precursor of the yeast mating pheromone alpha-factor and a synthetic sequence encoding a biologically active 24-amino acid carboxyl-terminal portion of the human atrial natriuretic peptide (hANP) precursor. Transformation of Saccharomyces cerevisiae with the hybrid gene resulted in the yeast cells secreting biologically active hANP into the extracellular medium. The secreted hANP was purified and found to be accurately processed at the junction in the chimeric alpha-factor/hANP protein, producing the desired mature hANP amino terminus. The secreted product was also folded correctly with respect to the single disulfide bond. However, the carboxyl terminus of the secreted hANP material was heterogeneous such that the major form lacked the last two amino acids of the peptide while the minor form was the full length material. The observed processing at the carboxyl terminus of the secreted hANP may reflect a normal processing event involved in alpha-factor peptide maturation.

Amino Acid Sequence↗

Effects of kinins on the isolated perfused rat kidney and evidence for the presence of renal B1 receptors.

The kinins, particularly lysyl-bradykinin (LBK), have a bimodal effect on the vasculature of isolated rat kidney. The vasorelaxant but not the vasoconstrictor effect of LBK seems to be mediated by prostaglandins. The vasoconstrictor action of LBK can be blocked by (L-Leu)8-des-Arg9-BK indicating that the rat kidney vasculature has B1 kinin-receptor which mediates the effects of kinins and/or their C-terminal metabolites.

Animals↗

Differing hemodynamic responses to atrial natriuretic factor in two models of hypertension.

Hemodynamic responses to synthetic atrial natriuretic factor (ANF), were studied in renin-dependent two-kidney, one-clip (2K,1C) and deoxycorticosterone (DOC) salt-treated hypertensive rats as well as normotensive controls. ANF infusion (800 pmol/kg prime, 120 pmol X kg-1 X min-1 for 60 min) decreased blood pressure (BP) more in conscious 2K,1C (-24 +/- 4%) than in DOC salt-treated (-12 +/- 4%, P less than 0.05) or control rats. Hemodynamic parameters were also evaluated during graded infusion of three doses, each for 30 min. At 24 and 120 pmol X kg-1 X min-1, ANF lowered BP in 2K,1C rats, both conscious (from 156 +/- 6 to 144 +/- 7, P less than 0.05 and 135 +/- 5 mmHg, P less than 0.05) and anesthetized (from 148 +/- 7 to 138 +/- 7, P less than 0.05 and 128 +/- 7, P less than 0.05). In anesthetized 2K,1C, BP changes were associated with reduction in total peripheral resistance (TPR) that became significant at 120 pmol X kg-1 X min-1 (-10 +/- 2%), whereas cardiac output (CO) and stroke volume (SV) were unchanged. In DOC-salt-treated rats these doses did not lower BP despite progressive falls in CO (-7 +/- 3% and -24 +/- 5%, P less than 0.05) and SV (-8 +/- 2% and -23 +/- 5%, P less than 0.05), which were balanced by a simultaneous rise in TPR (+12 +/- 4% and +26 +/- 10%, P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Relationship between renal hemodynamic and natriuretic effects of atrial natriuretic factor.

The degree by which atrial natriuretic factor (ANF)-induced renal hemodynamic changes account for its natriuretic effect was determined by early clamp experiments in six anesthetized dogs. After control periods, perfusion pressure of the left kidney (LK) was reduced to 80-90 mmHg, and synthetic ANF (auriculin A) was infused intravenously (0.3 micrograms X min-1 X kg body wt). After recovery, furosemide (F) was administered as a bolus injection (1 mg/kg body wt). In the right kidney (RK), which served as a time control, ANF increased (P less than 0.05) glomerular filtration rate (GFR) 16 +/- 4% and Na excretion (UNa V) 261 +/- 63%, whereas it decreased urine osmolality (Uosm) 36 +/- 7% without changing free water clearance. ANF also increased diuresis (V) and kaliuresis (UKV). F produced qualitatively the same results without changing GFR. In the clamped LK, ANF failed to increase GFR (22 +/- 4 vs. 26 +/- 4 ml/min), UNaV (30 +/- 9 vs. 33 +/- 11 mueq/min), V, and UKV or to decrease Uosm (841 +/- 97 vs. 840 +/- 114 mosmol/kg H2O). F had similar effects in LK as in RK. The data demonstrate that the natriuretic effect of ANF is abolished when its renal hemodynamic actions are impeded. In addition, the results demonstrate that ANF antagonizes renal vasoconstriction in the dog. The results are consistent with the view that the ANF-induced natriuresis is due in great part to an increase in the filtered load of Na into a washed-out inner medulla.

Animals↗

Cardiovascular effects of atrial natriuretic factor in anesthetized and conscious dogs.

Atrial natriuretic factor lowers blood pressure in normotensive and hypertensive animal models. The present study examined the mechanism of the blood pressure-lowering effect in 10 normotensive dogs. Four awake dogs previously instrumented with electromagnetic flow probes for measurement of cardiac output and catheters for systemic hemodynamic and cardiac dynamic measurements were studied. After a 30-minute control period, a 3 micrograms/kg bolus followed by 0.3 micrograms/min/kg of a 24-residue synthetic atrial natriuretic factor was infused for 30 minutes, followed by a 1-hour recovery period. Mean arterial pressure fell significantly during infusion (control, 125 +/- 4; infusion, 108 +/- 5; recovery, 125 +/- 9 mm Hg; p less than 0.05) and was accompanied by a slight but significant bradycardia (control, 144 +/- 7; infusion, 134 +/- 5; recovery, 145 +/- 7 beats/min; p less than 0.05). Significant reductions in cardiac output (control, 2.66 +/- 0.60; infusion, 2.18 +/- 0.60; recovery, 2.74 +/- 0.60 L/min; p less than 0.05), stroke volume (control, 18.4 +/- 3.9; infusion, 16.0 +/- 4.2; recovery, 19.0 +/- 3.7 ml/beat; p less than 0.05), and maximum increase in rate of change of left ventricular systolic pressure (control, 2475 +/- 200; infusion, 2088 +/- 216; recovery, 2487 +/- 243 mm Hg/sec; p less than 0.05) were also observed during infusion. No significant changes in total peripheral resistance or central venous pressure were noted, although the latter tended to fall during infusion. A similar pattern was observed in six pentobarbital-anesthetized dogs, except that infusion of atrial natriuretic factor did not induce bradycardia.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

[Structure and biological action of the atrial natriuretic factor (auriculin)].

The studies described in this report emphasize the potential importance of atrial natriuretic factor in affecting systemic and renal hemodynamics, sodium excretion and components of the renin-angiotensin-aldosterone system. It should be emphasized, however, that at present there is no direct and unequivocal evidence that ANF is secreted into the blood. Until this is done and until the factors that regulate its secretion have been established, its potential physiological role must remain speculative.

Adrenal Glands↗