Kallikrein has been localized in rodent kidney and salivary glands by means of an immunoglobulin-enzyme bridge technique. In sections of kidney, anti-kallikrein antibodies bound to the apical region of certain distal tubule segments in the cortex, to reabsorption droplets of proximal convoluted tubules, and to certain duct segments in the papilla. In salivary glands of both male and female rats and mice, and apical rim of most striated duct cells of submandibular, parotid and sublingual glands and granular tubules of submandibular glands exhibited immunoreactivity. Granular intercalated duct cells in female submandibular glands also displayed immunostaining for kallikrein. Phenylephrine administration resulted in loss of immunoreactive granules from the granular convoluted tubule cells of male mouse submandibular gland. This response was paralleled by a biochemically demonstrable decrease in kallikrein-like tosylarginine methyl ester (TAME) esterase activity.

The embryonic distribution of atriopeptin (atrial natriuretic factor) in the Sprague-Dawley rat heart was mapped by immunoperoxidase staining of embryonic and neonatal hearts using rabbit antiserum to atriopeptigen purified from adult rat atrium. During the period of cardiac septation (days 14 and 16), immune serum reacted strongly with myocardial cytoplasmic granules in two sites: the inner cell layer along the cephalic curvature of the atria and the trabeculae of the incompletely divided ventricles. The youngest hearts studied (gestational day 11) displayed only nonspecific diffuse peroxidase reactivity within blood cells, indistinguishable from control sections incubated with normal rabbit serum. One week following birth, intense anti-atriopeptin reactivity was widely distributed through both atria. In addition, immunoreactive cytoplasmic granules were found at several sites in the ventricular myocardium. Along the fiber tracts of the concentric layers of the ventricular walls and interventricular septum, scattered granular foci were seen between nuclei of contiguous elongated myocytes. Positive staining was also seen within the papillary muscles and trabeculae carnae, regions shown by Alcian blue/periodic acid-Schiff base staining of sister sections to be relatively rich in glycogen. These patterns of antibody reactivity suggest the coupling of early atriopeptin secretory activity with developing cardiac function.

Fixation of rat parotid with an unbuffered osmium tetroxide solution containing nearly saturated potassium (pyro)antimonate resulted in abundant deposition of cation-antimonate precipatates in acinar cells. Altering the antimonate concentration, including buffers or chelators in the solution or changing the primary fixative resulted in an altered intensity and distribution of the precipitates formed in the tissue, apparently reflecting a degree of selectivity in ion localization. Decreasing the concentration of pyroantimonate to about half-saturation preserved predominantly the less soluble antimonate salts (e.g., Na+, Ca++) and resulted in preferential retention of deposits along the plasmalemma and in mitochondrial "dense bodies," with loss of most cytoplasmic and nuclear precipitates. A similar pattern was seen if fixation with the high concentration antimonate-osmium procedure was followed by a prolonged rinse. Adding phosphate or collidine buffers markedly decreased precipitates in the nuclei and on granular reticulum as well. Phosphate buffer or ehtyleneglycoltetraacetate inhibited in vitro precipitation of calcium and sodium and decreased or abolished plasmalemmal deposits. Glutaraldehyde fixation, either in the presence of antimonate or prior to antimonate-containing osmium tetroxide, abolished heterochromatin deposits. Mitochondrial dense bodies were of two types, one containing precipitate and the other inherently osmiophilic. The latter were also observed in pyrophosphate-osmium controls. Results from in vitro titrations of cations with the various antimonate methods and from neutron activation analyses of fixed tissues supported conclusions drawn from fine structural distribution patterns and were interpreted as follows. In rat parotid acinar cells, deposits in heterochromatin and on granular reticulum probably arose from precipitation in sites of high K+ and H+ as well as--NH3+-rich histones. Plasmalemmal antimonate deposits demonstrated sites of sodium and/or calcium accumulation. Some mitochondrial dense bodies contained Ca++ whereas others were inherently osmiophilic. Large, extracellular deposits were probably predominantly sodium precipitates.

The organization and fine structure of granular convoluted tubule cells (GCT) from male mouse submandibular glands have been examined in controls and in animals injected with adrenergic and cholinergic secretagogues. Control submandibular glands exhibited a single population of GCT cells with numerous homogeneous granules filling the apical two-thirds of the cytoplasm. A zone of transition cells, exhibiting characteristics of both GCT and striated duct cells, was found between the agranular intercalated duct and GCT segments. These transition cells possessed apical granules of variable size as well as prominent basal striations. Dramatic changes in the morphology of GCT cells followed administration of the alpha-adrenergic agent, phenylephrine. The extensive degranulation involved formation of "secretory pools" of fused granules and release of secretory material into the lumen. The appearance of numerous smooth vesicles near luminal membranes suggested extensive membrane retrieval. Intracellular membrane-limited aggregates of membrane fragments suggested that much of the retrieved membrane was destined for degradation. Rough endoplasmic reticulum was highly dilated but there was no indication of increased size or activity of the Golgi complex. Ultrastructural evidence indicated that the secretory responses to isoproterenol, a beta-adrenergic agent, and to pilocarpine, a cholinergic agent, were much more modest, but it is clear that some secretory response to these agents does occur. The other cell types of the duct and tubule system did not exhibit comparable morphological changes in response to the agents used.

AbstructWe evaluated the blood pressure response to chrome salt loading m a rat stram inbred for low urmary kalhkrem excretion Low-kalhkrem rats showed greater systohc blood pressure values (130% 1 versus 114~2 mm Hg m controls, P< 05) at 9 weeks of age Systolic blood pressure was mcleased after 10 days of dietary sodmm loading m the low-kalhkrem group and remained unchanged m controls (1.53+1 versus 11222 mm Hg, P< 01) In additional experiments, blood pressure sensltlvlty to salt was tested m low-kalhkrem rats receiving a chronic infusion of rat glandular kalhkrem (1.7 pg/day per 100 g body weight, IV) or vehicle Systohc blood pressure of vehicle-treated rats was increased by salt loading (from 13821 to 15822, 153+1, and 145?2 mm Hg at 5, 10, and 15 days, respectively, P< 01), while It remained unchanged m the kalhkrem-treated group (from 136?2 to 146?5, 140+2, and 13424 mm Hg at 5, 10, and 15 days, respectively, P=NS) Urinary kalhkrem excrenon was m- sodnnn . kmms l blood pressure C osegregation of a sequence variation of the kalhkrem gene with high blood pressure phenotype in inbred strains derived from the spontaneously hypertensive rat and the normotenslve Brown Norway rat suggests that this gene may have the capacity to affect blood pressure. 1 This hypothesis 1s supported by the observation that kinms, which are derived from the enzymatic action of kalhkrem on kmmogen, cause vasodilation, dmresls, and natrmresis. Urinary kalhkrem excretion 1s reduced m genetic hypertension. In particular, the lowkallikrem phenotype has been used as a marker of salt Sensitivity in normotensive SUbJeCtS as well as m essential hypertensive patients 2.3 These studies also suggest that depressed activity of the renal kalllkrem-kmm system could contribute m the pathogenesis of salt-dependent hypertension Recently, a stram was developed m our laboratory, by breeding rats from a Wlstar stock accordmg to urmary kalhkrem phenotype, exclusively 4 This breeding approach represents an important difference compared with other experlmental models of genetic hypertension m which urmary kalllkrem excretion has been found to be decreasede5-7 Indeed, the low-kalhkrem phenotype could be secondary to the increase m blood pressure m the latter strains We found that application of chetary sodmm load- Correspondence to Paolo Madeddu, MD, Chmca Mechca, Umversity of Sassarl, Viale S Pietro 8, 07100 Sassan, Italy E-mad madeddu@mbox vol it 0 1997 Amencan Heart Association, Inc mg induces a significant increase m the blood pressure of low-kalhkrem rats compared with normal-kalhkrem Wistar rats, an effect that was attributed to sodium retention.4 These results mdlcate an association between a genetltally determined defect in urinary kalhkrem excretion and blood pressure sensitivity to salt The aim of the present study was to evaluate the effect of rat glandular (or tissue) kalhkrem admmistratlon on the exaggerated blood pressure response to salt m the low-kalhkrem rats In addition, Smce ludney weight was found to be reduced m this strain, we wished ...