Effect of formalin fixation on the dry mass of isolated rat liver nuclei

Summary Effect of formalin fixation on the dry mass of isolated rat liver nuclei was studied by interference microscopy. It was found that a fixation time of 2–6 hrs prevents completely the elution of water-extractable material from nuclei. Nuclei unfixed, water-extracted for 4 hrs lose about 20% of their initial dry mass. Fixation time longer than 6 hrs resulted also in a decrease of dry mass of the nuclei. Liquid scintillation counting of ¹⁴C-formaldehyde served for finding the influence of bound formaldehyde on dry mass values. As calculated, after 2 hrs fixation with formalin the bound formaldehyde constitutes 3.6% of the dry mass of isolated untreated nuclei.     

Rate of protein synthesis in rat salivary gland cells after pilocarpine or feeding

In untreated, fasting animals the cells of the serous demilunes of the sublingual gland incorporate [3H]-leucine at a higher rate than any other of the 5 main cell types of the 3 major salivary glands. The acinar cells of the submandibular and the mucous cells of the sublingual gland show intermediate values, while the cells of the granular ducts of the submandibular and the acini of the parotid gland have a low rate of incorporation. In fasting animals extrusion of newly synthesized protein starts early in the cells of the serous demilunes. It starts between 4 and 7 hrs after [3H]-leucine injection in the acinar cells of the submandibular gland, while the other cell types did not lose substantial amounts of labelled (glyco)protein within 7 hrs. The secretion of protein is stimulated by the cholinergic drug pilocarpine in all but one of the 5 types of salivary gland cells studied. The acinar cells of the submandibular gland react strongly, the granular duct cells less strongly. Still less are the reactions of the acinar cells of the parotid and of the nucous cells of the sublingual gland. The cells of the serous demilunes of the latter appear to be insensible to pilocarpine. The effect of food uptake on secretion does not differ from pilocarpine stimulation, with one exception: the acinar cells of the parotid gland react more strongly on food uptake than on cholenergic stimulation.     

Nuclear changes in allium cepa in response to infection by fusarium moniliforme

Fusarium moniliforme was isolated [into culture] from white onion and inoculated into holes bored in onion bulbs. After 48 hrs, the nuclear dry mass of onion cells 0–5 mm ahead of the mycelium had decreased by 19% as determined by quantitative interference microscopy as compared to nuclear dry mass decreases of 11 % caused by avirulent Aspergillus niger, 26 % by an unidentified onion pathogen, 41 % by virulent Botrytis allii, and 42 % by virulent Aspergillus niger. In the present study, the nucleolar dry mass declined by 34 % in response to F. moniliforme under the same conditions. Degradation or exit of nuclear and nucleolar macromolecules in response to fungal products secreted in advance of mycelium may be responsible for lower nuclear and nucleolar dry mass.     

Interferenzmikroskopische und cytophotometrische Untersuchungen zur Analyse von Zellkernen

Zusammenfassung In vergleichenden interferenzmikroskopischen und cytophotometrischen Untersuchungen wurde versucht, die Globulinfraktion von Zellkernen quantitativ zu erfassen. Dazu wurden Thymuslymphozyten und nach Behrens isolierte Leberzellkerne der Ratte und Nierenzellkerne vom Schwein im Ausstrichpräparat bis zu 30 Std in physiologischer Kochsalz- und Tyrodelösung extrahiert. Die Abnahme der Kerntrockenmasse wurde interferenzmikroskopisch gemessen und der DNS-, Histon- und Gesamtproteingehalt cytophotometrisch bestimmt (Feulgen-Reaktion, Färbung mit Gallocyanin-Chromalaun, Fastgreen bei pH 8 und pH 2, Naphtholgelb S).Bei Verwendung von Glycerin als Eindeckmittel fand sich bei Leberzellkernen nach Extraktion mit 0,14 m NaCl-Lösung von 3–5°C eine Trockengewichtsabnahme von 18% nach 2 Std, von 29% nach 10 Std und von 34% nach 30 Std. Bis zu 2 Std wurden Proteine, nach 10 Std Nukleohistone extrahiert. Da der nach 2 Std gemessene Proteinverlust gut mit dem Globulingehalt von Leberzellkernen übereinstimmt, wurde offenbar die Globulinfraktion erfaßt. Nach Extraktion mit Tyrodelösung bei Zimmertemperatur lag der Substanzverlust in der gleichen Größenordnung (2 Std: 12%, 10 Std: 20%, 30 Std: 29%).Bei Verwendung von Tyrodelösung als Eindeckmittel blieb nach Extraktion isolierter Nierenzellkerne mit 0,14 mNaCl-Lösung die Kerntrockenmasse bis zu 2 Std unverändert, da die Globulinfraktion schon während der interferenzmikroskopischen Messung in Lösung geht. Auch bei Thymuslymphozyten fand sich erst nach 24 Std ein Verlust von 18% der Kerntrockenmasse durch Extraktion von DNS und Histonen.Während der Suspension von Thymusgewebe in 0,14 mNaCl-Lösung nahm innerhalb von 4 Std das Kerntrockengewicht um 40% zu, wahrscheinlich infolge Proteinaufnahme.Die gewonnenen Ergebnisse werden hinsichtlich der Fehlerquellen und im Vergleich zu cytophotometrischen Messungen und biochemischen Globulinbestimmungen diskutiert.

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Analysis of cell nuclei with the interference-microscope and cytophotometryI. Determination of globulins

Summary The globulinfraction of cell nuclei was determined by dry mass determinations in comparison with cytophotometric measurements.Lymphocytes from the thymus gland, and liver nuclei from the rat and kidney nuclei from the pig, isolated with Behrens' technique, were extracted in smears, up to 30 hrs, in physiological saline and tyrode solution. The decrease of nuclear dry mass was measured with the interference microscope, and the content of DNA, histones and total proteins was determined by cytophotometry (Feulgen-reaction, staining with gallocyanin chrome alum, fast green at pH 8 and pH 2, naphtol yellow S).Using glycerol as embedding medium, after extraction with 0,14 M saline at 3—5° C the dry weight of liver nuclei decreased by 18% after 2 hrs, by 29% after 10 hrs and by 34% after 30 hrs. Nuclear proteins were extracted up to 2 hrs, nucleo-histones were removed after 10 hrs. Since the loss of proteins is in good agreement with the globulin content of liver nuclei, obviously the nuclear globulins have been extracted until 2 hrs. After extraction with tyrode solution at room temperatue, the loss of dry matter was in the same range (2 hrs: 12%, 10 hrs: 20%, 30 hrs: 29%).Using tyrode solution as embedding medium, the dry weight of kidney nuclei remained unchanged for 2 hrs when extracted with 0,14 M saline, since the globulin-fraction is lost while nuclei are measured by interferometry. With thymocytes only a loss of 18% of nuclear dry weight was found after 24 hrs due to removal of DNA and histones.Suspending the thymus gland in 0,14 M saline, the nuclear dry weight increased by 40% in 4 hrs, probably due to adsorption of proteins.The results were discussed in the light of the errors of the measurements and compared to biochemical and histochemical datas.

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Mit Unterstützung durch die Deutsche Forschungsgemeinschaft.     

How much pilocarpine contaminates pilocarpine‐induced tick saliva?

Pilocarpine is often applied or injected into ticks to induce salivation, and the resulting saliva used to test for various pharmacological, biochemical and immunological activities. To measure the amount of pilocarpine in pilocarpine-induced tick saliva, an HPLC-MS/MS method, based on capillary strong cation exchange chromatography online with an ion trap mass spectrometer, was used to measure pilocarpine in the pg to ng range. Results indicate large concentrations of pilocarpine in Ixodes scapularis Say and Amblyomma americanum (Linnaeus) (Acari: Ixodidae) saliva, ranging from 3 to 50 mm. Due to the known effects of pilocarpine on smooth muscle and immune cells, appropriate controls are proposed and discussed for proper interpretation of results using this saliva preparation.     

Production of imidazole alkaloids in cell cultures of jaborandi as affected by the medium pH

The effect of pH (from 4.8 to 9.8) on the production of pilosine and pilocarpine and on their partition between cell and medium was studied in two lineages (P and PP) of Pilocarpus microphyllus cell suspension cultures. Highest mass accumulation was observed at high pHs and both lineages produced pilocarpine while only lineage PP produced pilosine. Both alkaloids were released in the medium but higher accumulation occurred in the cells. The highest production of pilocarpine was at pH 8.8–9.8 in both cell lineages. Other imidazole alkaloids were also identified in both lineages. At all pHs tested, the pH in the media cultures tended to stabilize around 6 after 10–15 days of cultivation. NO₃ ⁻ and NH₄ ⁺ variation in the media might partially explain the pH stabilization.     

Further studies on the effect of cimetidine and other neurotropic drugs on rat serum prolactin

The hyperprolactinemic effect of H2 histamine receptor antagonists has been described in the rat and in humans. The present study was undertaken to explore more fully the hyperprolactinemic action of cimetidine, and its interrelationship with other neurotropic agents. Adult ovariectomized estrogen-primed rats were injected with mepiramine, diphenhydramine, pilocarpine, atropine, dopamine, cimetidine or saline as control, at different sequences and the effect on serum prolactin was determined. The effect of cimetidine on prolactin release "in vitro" by hemipituitaries of estrogenized male rats during a short time incubation period, was also investigated. Our results indicate that cimetidine is able to release prolactin and that this effect is not prevented by atropine or the classical antihistaminergic agents mepiramine and diphenhydramine. Both pilocarpine and dopamine inhibit the prolactin release due to cimetidine. The hypoprolactinemic action of pilocarpine was completely blocked by atropine, but not by mepiramine or diphenhydramine. Finally, cimetidine was unable to modify significantly the prolactin release by incubated pituitaries. It is postulated that cimetidine acts mainly at the brain and that the hyperprolactinemic effect is not mediated by muscarinic or H1 histaminergic receptors. This action can be prevented by drugs such as dopamine that are able to act on the lactotroph, and also by pilocarpine, probably by stimulating a prolactin inhibiting pathway.     

Significance of saliva for the denture‐wearing population

This paper summarises a series of studies already published in German and presents new data related to the aetiology of the dry mouth' and its associated problems. Aims: to study factors affecting mucous and serous salivary gland secretion, the aetiology of the ‘dry mouth’ and its associated problems, causative factors for hyposalivation and it's treatment Setting : two university dental hospitals. Subjects: 587 denture wearers and 521 control subjects, and autopsy material Interventions : exercise, chewing, water, oestrogen, pilocarpine, and anetholtrithion theiapy, biopsy of the minor glands Main outcome measures : Palatal secretion (PAL, μL/cm²/min) and parotid salivary flow (PAR), subjective complaints and clinical findings. Results: resting flow rates for PAL between 0 and 65 μl/cm²/min were seen in every age group. The flow rates of PAR (0 to 3.7 ml/10 min) were not correlated with PAL. Most patients with a resting flow rate of PAL≤6.0 μl/cm² suffer from a ‘dry mouth’ and Burning Mouth Syndrome (BMS) or oral dysaesthesia (OD) with or without chronic lesions of the oral mucosa. Etiological factors for the incidence of reduced PAL and associated problems include xerostomic drugs, oestrogen deficiency, ladiotherapy, thyroid dysfunction, smoking or continuous wearing of complete upper dentures. PAL also correlated with the retention of upper complete dentures. PAL was correlated with the water content of epithelial tissues. PAL and PAR were both increased by drinking ample fluid, improving their circulation by physical exercises, chewing intensively, or taking oestrogens, pilocarpine, anetholtrithion. Conclusions: Variation in palatal salivary secretion occurs and is clinically important.     

Studies on the relationship between the oxygen uptake and the release of amylase and sialic acid

The relationship between the oxygen uptake and the release of amylase and sialic acid induced by pilocarpine was investigated in dog submandibular glands. Pilocarpine dose-dependently stimulated the oxygen uptake. The dose required for the maximal response was 10 microM. The release of amylase and sialic acid induced by pilocarpine was inhibited by the addition of iodoacetic acid, malonic acid, 2, 4-dinitrophenol, antimycin A or sodium azide. The oxygen uptake induced by pilocarpine was significantly inhibited by iodoacetic acid, malonic acid, antimycin A or sodium azide. On the other hand, 2, 4-dinitrophenol further stimulated the oxygen uptake by pilocarpine. The increase in the oxygen uptake or the release of amylase and sialic acid induced by pilocarpine was significantly inhibited by ouabain. The Na+, K+-ATPase activity ratio in the microsomal fraction of dog submandibular glands was dose-dependently increased by pilocarpine. The Na+, K+-ATPase activity ratio induced by pilocarpine was significantly inhibited by ouabain, antimycin A, oligomycin or 2, 4-dinitrophenol. The pilocarpine-induced Na+, K+-ATPase activity ratio was significantly inhibited by the removal Ca2+ from the medium or the addition of 2 mM EGTA. These results suggest that the increase in the oxygen uptake by pilocarpine is profoundly involved in the energy supply for the process of amylase and sialic acid release. In particular, the energy supply demanded for the activation of Na+ pump may play a role in the mechanism by which pilocarpine induces the oxygen uptake.     

Efficiency of induced mutagenesis at early stages (gametes,zygotes, proembryos) of ontogenesis in Nigella damascena L.

Summary An original technique has been developed of treating gametes, zygotes and early embryos of Nigella damascena L. with chemical and physical mutagens. A delay in fertilization and a decrease in the rate of cell division of the embryo and the endosperm after mutagen treatment have been found. Our method of treating gametes, zygotes and proembryos with chemical and physical mutagens is, by all criteria, superior to that of treating dry seeds. Treatment applied at early stages of ontogenesis not only induced a much higher mutation ratio compared with dry seeds, but also gave a broader mutation spectrum. The 55 types of hereditary change obtained affect the structure of vegetative and reproductive organs. Mutations which change the structure of the reproductive organs of flowers are of specific interest.The optimum dose for this object and the method of treatment which induces high mutation ratio (up to 96 % of families with changes) is 0.003p (16 hrs) for ethylenimine and 0.005p or 0.008p (16 hrs) for nitrosomethylurea. Treatment of dry seeds turned out to be much less effective.     

Differences in sensitivity to the convulsant pilocarpine in substrains and sublines of C57BL/6 mice

In rodents, the cholinomimetic convulsant pilocarpine is widely used to induce status epilepticus (SE), followed by hippocampal damage and spontaneous recurrent seizures, resembling temporal lobe epilepsy. This model has initially been described in rats, but is increasingly used in mice, including the C57BL/6 (B6) inbred strain. In the present study, we compared the effects of pilocarpine in three B6 substrains (B6JOla, B6NHsd and B6NCrl) that were previously reported to differ in several behavioral and genetic aspects. In B6JOla and B6NHsd, only a small percentage of mice developed SE independently of whether pilocarpine was administered at high bolus doses or with a ramping up dosing protocol, but mortality was high. The reverse was true in B6NCrl, in which a high percentage of mice developed SE, but mortality was much lower compared to the other substrains. However, in subsequent experiments with B6NCrl mice, striking differences in SE induction and mortality were found in sublines of this substrain coming from different barrier rooms of the same vendor. In B6NCrl from Barrier #8, administration of pilocarpine resulted in a high percentage of mice developing SE, but mortality was low, whereas the opposite was found in B6NCrl mice from four other barriers of the same vendor. The analysis of F1 mice from a cross of female Barrier 8 pilocarpine‐susceptible mice with resistant male mice from another barrier (#9) revealed that F1 male mice were significantly more sensitive to pilocarpine than the resistant parental male mice whereas female F1 mice were not significantly different from resistant Barrier 9 females. These observations strongly indicate X‐chromosome linked genetic variation as the cause of the observed phenotypic alterations. To our knowledge, this is the first report which demonstrates that not only the specific B6 substrain but also sublines derived from the same substrain may markedly differ in their response to convulsants such as pilocarpine. As the described differences have a genetic basis, they offer a unique opportunity to identify the genes and pathways involved and contribute to a better understanding of the underlying molecular mechanisms of seizure susceptibility.     

Intracellular Water Exchange for Measuring the Dry Mass,Water Mass and Changes in Chemical Composition of Living Cells

We present a method for direct non-optical quantification of dry mass, dry density and water mass of single living cells in suspension. Dry mass and dry density are obtained simultaneously by measuring a cell’s buoyant mass sequentially in an H₂O-based fluid and a D₂O-based fluid. Rapid exchange of intracellular H₂O for D₂O renders the cell’s water content neutrally buoyant in both measurements, and thus the paired measurements yield the mass and density of the cell’s dry material alone. Utilizing this same property of rapid water exchange, we also demonstrate the quantification of intracellular water mass. In a population of E. coli, we paired these measurements to estimate the percent dry weight by mass and volume. We then focused on cellular dry density – the average density of all cellular biomolecules, weighted by their relative abundances. Given that densities vary across biomolecule types (RNA, DNA, protein), we investigated whether we could detect changes in biomolecular composition in bacteria, fungi, and mammalian cells. In E. coli, and S. cerevisiae, dry density increases from stationary to exponential phase, consistent with previously known increases in the RNA/protein ratio from up-regulated ribosome production. For mammalian cells, changes in growth conditions cause substantial shifts in dry density, suggesting concurrent changes in the protein, nucleic acid and lipid content of the cell.     

Effects of sialagogues on the syntheses of polyamines and DNA in murine parotid gland

When rat parotid explants were cultured on siliconized lens paper floating on chemically defined 199 medium, all of sialagogues tested increased ornithine decarboxylase activity, which was roughly proportional to the amylase released into the culture medium. S-Adenosylmethionine decarboxylase and DNA synthesis were also induced by isoproterenol, methoxamine, carbachol and pilocarpine, but not by serotonin or substance P. The increases of the two decarboxylase activities and DNA synthesis were observed in vivo in mouse parotid gland after repeated injections of carbachol or pilocarpine. These results indicate that both adrenergic and cholinergic sialagogues stimulate the syntheses of polyamines and DNA in murine parotid gland.     

Mechanisms of organophosphate insecticide-induced airway hyperreactivity

It has been suggested that pesticide exposure may be a contributing factor underlying the increased incidence of asthma in the United States and other industrialized nations. To test this hypothesis, airway hyperreactivity was measured in guinea pigs exposed to chlorpyrifos, a widely used organophosphate pesticide. Electrical stimulation of the vagus nerves caused frequency-dependent bronchoconstriction that was significantly potentiated in animals 24 h or 7 days after a single subcutaneous injection of either 390 mg/kg or 70 mg/kg of chlorpyrifos, respectively. Mechanisms by which chlorpyrifos may cause airway hyperreactivity include inhibition of acetylcholinesterase (AChE) or dysfunction of M3 muscarinic receptors on airway smooth muscle or of autoinhibitory M2 muscarinic receptors on parasympathetic nerves in the lung. AChE activity in the lung was significantly inhibited 24 h after treatment with 390 mg/kg of chlorpyrifos, but not 7 days after injection of 70 mg/kg of chlorpyrifos. Acute exposure to eserine (250 microg/ml) also significantly inhibited lung AChE but did not potentiate vagally induced bronchoconstriction. Neuronal M2 receptor function was tested using the M2 agonist pilocarpine, which inhibits vagally induced bronchoconstriction in control animals. In chlorpyrifos-treated animals, pilocarpine dose-response curves were shifted significantly to the right, demonstrating decreased responsiveness of neuronal M2 receptors. In contrast, chlorpyrifos treatment did not alter methacholine-induced bronchoconstriction, suggesting that chlorpyrifos does not alter M3 muscarinic receptor function on airway smooth muscle. These data demonstrate that organophosphate insecticides can cause airway hyperreactivity in the absence of AChE inhibition by decreasing neuronal M2 receptor function.     

Estrogen regulation of the rat anterior pituitary gland proteome

Estrogen is known to affect the regulation of all six of the established anterior pituitary gland (AP) hormones, but little is known of the specifics of its regulation of the AP hormones, their isoforms, and nonhormonal AP proteins. We used difference gel electrophoresis in conjunction with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and peptide mass fingerprinting to quantify the effects of estrogen on the AP-soluble protein fraction in rats. Two-month-old rats were ovariectomized and used at 6 months of age. They were injected subcutaneously with sesame oil vehicle or 50 mug estradiol valerate in vehicle and studied 48 hrs later, approximately 3 hrs before the time of the anticipated onset of the estrogen-induced surges of gonadotropins in blood. The APs were pooled, and the soluble protein fraction was examined in replicate analyses. After DeCyder software analysis, we identified 26 protein spots that had a 1.5-fold or greater average increase in the experimental group relative to the controls. Nineteen showed a 1.5-fold or greater decrease. Estrogen increased levels of the more acidic isoforms of growth hormone and prolactin and of proteins involved in protein synthesis, folding, and secretion (e.g., eukaryotic translation elongation factor 2, ERp57, ERp29, Hsc70-ps1, calreticulin, coatomer delta subunit, and secretogranin II) and of some metabolic enzymes (e.g., arginosuccinate synthetase, enolase 1, creatine kinase B, phosphoglycerate mutase, malate dehydrogenase, pyruvate kinase, and aldolase A). The majority of the downregulated proteins were involved in RNA or DNA interactions (e.g., five heterogeneous nuclear ribonucleoproteins, DEAD-box proteins 17 and 48, ssDNA binding protein PUR-alpha, PTB-associated splicing factor, and Pigpen protein), but isovaleryl coenzyme A dehydrogenase, mitochondrial aldehyde dehydrogenase, stathmin 1, vinculin, radixin, and secretogranin III were also reduced. Our results indicate that estrogen acts in vivo within 48 hrs to modulate levels of a significant number of AP proteins.     

HMG-CoA reductase inhibitor decreases small dense low-density lipoprotein and remnant-like particle cholesterol in patients with type-2 diabetes

Our studies have focused on the effect of injection of L-NAME and sodium nitroprussiate (SNP) on the salivary secretion, arterial blood pressure, sodium excretion and urinary volume induced by pilocarpine which was injected into the medial septal area (MSA). Rats were anesthetized with urethane (1.25 g/kg b. wt.) and a stainless steel cannula was implanted into their MSA. The amount of saliva secretion was studied over a five-minute period after injection of pilocarpine into MSA. Injection of pilocarpine (10, 20, 40, 80, 160 microg/microl) into MSA produced a dose-dependent increase in salivary secretion. L-NG-nitro arginine methyl-esther (L-NAME) (40 microg/microl), a nitric oxide (NO) synthase inhibitor, was injected into MSA prior to the injection of pilocarpine into MSA, producing an increase in salivary secretion due to the effect of pilocarpine. Sodium nitroprussiate (SNP) (30 microg/microl) was injected into MSA prior to the injection of pilocarpine into MSA attenuating the increase in salivary secretion induced by pilocarpine. Medial arterial pressure (MAP) increase after injections of pilocarpine into the MSA. L-NAME injected into the MSA prior to injection of pilocarpine into MSA increased the MAP. SNP injected into the MSA prior to pilocarpine attenuated the effect of pilocarpine on MAP. Pilocarpine (40 ug/ul) injected into the MAS induced an increase in sodium and urinary excretion. L-NAME injected prior to pilocarpine into the MSA increased the urinary sodium excretion and urinary volume induced by pilocarpine. SNP injected prior to pilocarpine into the MSA decreased the sodium excretion and urinary volume induced by pilocarpine. All these roles of pilocarpine depend on the release of nitric oxide into the MSA. We may also conclude that the MSA is involved with the cholinergic excitatory mechanism that induce salivary secretion, increase in MAP and increase in sodium excretion and urinary volume.     

Disjunction between the daily duration of light and the plasma level of prolactin under hyperlong days in rams

Adult rams were submitted to an artificial light regime with increasing and decreasing daylengths between 6 and 20 hrs. over a period of 8 months. Blood samples occurred at different durations of daylengths for measuring plasma levels of prolactin. Levels were high for a daily duration of light of 14.20, 17 and 18.30 hrs. along the increasing light-phase and of 18.30 hrs. during the decreasing light-phase. On the other hand prolactin levels were low for short daylengths, but also for a daylength of 20 hrs. It clearly appears that hyperlong days (greater than 18 hrs.) do not stimulate (or do actively inhibit) the secretion of prolactin in ovines.     

Troop movement and food habits of white-faced monkeys in a tropical-dry forest

Home-range, movement and activity patterns and diet of a group of 16 white-faced monkeys (Cebus capucinus) were studied in a Costa Rican tropical dry forest at the end of the dry season (March and April) of 1982. The troop used an area of .67 km2 and moved an average of 4.5 km +/- 0.6 daily. Three basic daily activities were identified: resting, moving and feeding. Resting was greatest between 1000-1500 hours. Movement, which was strongly correlated with feeding, occurred most between 0500-0900 hrs and 1300-1800 hrs. The primates were omnivores, feeding on parts of 27 species of plants, four species of insects and an Anolis lizard.     

Nitric oxide of the supraoptic nucleus influences the salivary secretion, sodium renal excretion, urinary volume and arterial blood pressure induced by pilocarpine

Male Holtzman rats weighting 200-250 g were anesthetized with zoletil 50 mg/Kg (tiletamine chloridrate 125.0 mg and zolazepan chloridrate 125.0 mg) into quadriceps muscle and stainless steel cannulas were implanted into their supraoptic nucleus (SON). We investigated the effects of the injection into the supraoptic nucleus (SON) of FK 409, a nitric oxide donor, and NW-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase inhibitor (NOS), on the salivary secretion, arterial blood pressure, sodium excretion and urinary volume induced by pilocarpine, which was injected into SON. The drugs were injected in 0.5 microl volume over 30-60 s. Controls was injected with a similar volume of 0.15 M NaCl. FK 409 and L-NAME were injected at doses of 20 microg/0.5 microl and 40 microg/0.5 microl respectively. The amount of saliva secretion was studied over a five-minute period after injection of pilocarpine into SON. Injection of pilocarpine (10, 20, 40, 80, 160 microg/microl) into SON produced a dose-dependent increase in salivary secretion. L-NAME was injected into SON prior to the injection of pilocarpine into SON, producing an increase in salivary secretion due to the effect of pilocarpine. FK 409 injected into SON attenuating the increase in salivary secretion induced by pilocarpine. Mean arterial pressure (MAP) increase after injections of pilocarpine into the SON. L-NAME injected into the SON prior to injection of pilocarpine into SON increased the MAP. FK 409 injected into the SON prior to pilocarpine attenuated the effect of pilocarpine on MAP. Pilocarpine (0.5 micromol/0.5 microl) injected into the SON induced an increase in sodium and urinary excretion. L-NAME injected prior to pilocarpine into the SON increased the urinary sodium excretion and urinary volume induced by pilocarpine. FK 409 injected prior to pilocarpine into the SON decreased the sodium excretion and urinary volume induced by pilocarpine. All these roles of pilocarpine depend on the release of nitric oxide into the SON. In summary the present results show: a) SON is involved in pilocarpine-induced salivation; b) that mechanism involves increase in MAP, sodium excretion and urinary volume.