Effect of chronic intermittent immobilization stress on Fos-like immunoreactivity in rat brain and adrenal medulla.

The present study examined the influence of short- and long-term chronic intermittent immobilization stress throughout the brain and on the adrenal medulla of intact rats using Fos-like immunoreactivity (Fos-LI) as a marker of cellular activation. The effect of adreno-medullectomy on the central nervous system (CNS) response to chronic immobilization stress was also examined. It was found that control unoperated, unstressed rats had no Fos-LI cells in the brain or in the adrenal medulla. In intact rats, neither short term (1 week) nor long term (4 weeks) chronic intermittent immobilization stress produced significant increases in Fos-LI in the CNS compared with control animals. However, marked increase in the number of Fos-LI cells was observed in the adrenal medulla of animals stressed for 4 weeks compared with control, unstressed animals or those stressed for 1 or 2 weeks. In adreno-medullectomised rats, 4 weeks, but not 1 week, chronic immobilization stress produced significant increases in numbers of Fos-LI neurons in the paraventricular hypothalamic and supraoptic nuclei and the medial amygdala compared with intact animals stressed for a similar period of time. It is concluded that long term stress produces chronic Fos-LI in the adrenal medulla and that adreno-medullectomy increases the Fos response of the PVN, supraoptic nucleus and medial amygdala to long term stress.

Reduction of plasma catecholamines in humans during clinically controlled severe underfeeding.

BACKGROUND: Sympathetic hyperactivity is one factor for alterations encountered in the plurimetabolic syndrome, a cluster of metabolic abnormalities including obesity, hyperlipidemia, sometimes hyperglycaemia, and hypertonia. It was interesting to know if prolonged severe underfeeding (230 kcal/day) leads to decreases in catecholamines in those patients. METHODS: The plasma concentrations of catecholamines in patients (n = 16) suffering from plurimetabolic syndrome were studied before and during a 16-day period of medically controlled severe underfeeding (230 kcal/day) at rest and in response to exercise. RESULTS: During the period of underfeeding, mean norepinephrine concentrations decreased at rest from 1.45 to 0. 96 nmol/liter, and in response to exercise, from 6.1 to 3.2 nmol/liter. Epinephrine concentrations decreased from 0.15 to 0.1 nmol/liter and from 0.26 to 0.17 nmol/liter, respectively. A significant decrease in catecholamine concentrations was observed only after 16 days of underfeeding. CONCLUSIONS: Clinically controlled underfeeding of patients with plurimetabolic syndrome may result in beneficial clinical and biochemical effects. The findings indicate that relatively long periods of underfeeding induce decreases in plasma catecholamine concentrations. Nevertheless, most of the fall in mean values in norepinephrine and also of the fall in blood pressure values occurred by Day 2. From those tendencies and from the significant changes in both parameters at Day 16 of severe underfeeding one could conclude that altered sympathetic nervous system activity could contribute to the fall in blood pressure.

Differential expression of SNAP-25 isoforms and SNAP-23 in the adrenal gland.

In the rat adrenal gland, we previously observed that SNAP-25 is not restricted to the plasmalemma in noradrenergic cells as it is in adrenergic cells, and hypothesized that SNAP-25 isoform expression is different in the two phenotypes. Expression of SNAP-25 isoforms and SNAP-23 was examined by immunoblotting, immunofluorescence, and RT-PCR. Amplifications of SNAP-25 mRNAs were combined with Southern hybridization, restriction enzyme analysis, and sequencing of cloned PCR products to compare SNAP-25 isoform expression in rat and bovine adrenal glands. SNAP-25 and SNAP-23 mRNA and protein are expressed in the glands; SNAP-23 is enriched in the adrenal cortex, whereas SNAP-25 is restricted to the adrenal medulla. Furthermore, high levels of SNAP-25 and low levels of SNAP-23 are observed in the PC12 cells, whereas both SNAP-25 and SNAP-23 are expressed in adrenal medullary cultures. In all extracts, the SNAP-23 mRNA corresponded to SNAP-23a. SNAP-25a is the major form expressed in rat adrenal glands (75%), as it is in PC12 cells (80%), but both SNAP-25a and SNAP-25b (40% vs. 60%) are expressed in bovine adrenal medulla in situ and in culture. In addition, an enriched population of adrenergic cells (93%) expressed a higher level of SNAP-25b (70%), suggesting that this isoform may not be restricted to fast neurotransmission.

[Peptide mediators in adrenal chromaffin cells: regulation of catecholamine selective secretion]. / Peptidnye mediatory v khromaffinnykh kletkakh nadpochechnikov: reguliatsiia izbiratel'noi sekretsii katekholaminov.

The data obtained suggest a potential mechanism that may account for the selective control of adrenaline and noradrenaline release from adrenal chromaffin cells. Some neuropeptides seem to affect in a different way the release from A- and NA-adrenal cells by means of regulating a set of cytochemical events: specific reception of cholinergic transmitters, expression of the second messenger system including cGMP and changes in Ca channels activity, changes in the catecholamine biosynthesis in adrenal chromaffin cells. Modulating function of substance P, endothelins, PACAP, and ANF, is discussed.

Substance P and epibatidine-evoked catecholamine release from fractionated chromaffin cells.

Bovine chromaffin cells were separated by density gradient centrifugation into subfractions enriched with either > 90% adrenaline- or 70-80% noradrenaline-producing cells. Concentrations of epibatidine (an alkaloid with nicotinic receptor activity) as low as 10 nM released adrenaline and noradrenaline from both fractions of cells maintained as monolayer cultures. The maximal effect was evoked by 30 nM epibatidine and was comparable to that evoked by 10 microM nicotine. The catecholamine release from the noradrenaline fraction was 30-40% higher than from the adrenaline fraction. Initial exposure to 50 nM epibatidine reduced release induced by a second exposure to the drug. There was cross-desensitization between epibatidine and nicotine. Substance P inhibited the epibatidine-evoked catecholamine release from both fractions by up to 85% (IC50 = 3-5 microM). The release of noradrenaline was inhibited more than that of adrenaline. In addition, substance P protected the chromaffin cells against desensitization of the nicotinic receptor by epibatidine. The C-terminal heptapeptide sequence of substance P was 10 x less active, two N-terminal sequences did not modulate the catecholamine release.

Catecholamine release from fractionated chromaffin cells.

Bovine chromaffin cells were separated by density gradient centrifugation into subfractions. After centrifugation on a self-generating Percoll gradient (42.75% isotonic Percoll, 30,000 x g for 22 min at 20 degrees C), the chromaffin cells were found in two clearly distinguishable peaks. The peak with the lower density contained most of the noradrenaline-producing cells (approximately 80%), whereas the adrenaline-producing cells were equally distributed between the two peaks. After collection of suitable fractions from the gradient, cell cultures were obtained, which were enriched with either > 90% adrenaline- or approximately 65% noradrenaline-producing cells. When stimulated by nicotine or carbachol, the dose-response curves of both cell fractions yielded similar EC50s for the release of adrenaline and noradrenaline. On the other hand, the cells of the less dense fraction released 30% more catecholamines (adrenaline as well as noradrenaline) than the cells of the more dense fraction. It is suggested that there are subpopulations among the adrenaline- and noradrenaline-producing cells with differences in receptor-effector coupling.

Functional studies with substance P analogues: effects of N-terminal, C-terminal, and C-terminus-extended analogues of substance P on nicotine-induced secretion and desensitization in cultured bovine adrenal chromaffin cells.

Substance P (SP) and SP analogues, including C-terminal, N-terminal, and C-terminus-extended analogues, have been investigated for their ability to modulate nicotine-induced secretion from bovine adrenal chromaffin cells in culture. Secretion was monitored by measuring the release of endogenous catecholamines by electrochemical detection following separation on HPLC and the release of endogenous ATP with an on-line luciferin-luciferase bioluminescence technique. SP is known to have the following two effects on nicotine-induced secretion of catecholamines (see Livett and Zhou, 1991): inhibition of the nicotinic response and protection against nicotinic desensitization. Secretion induced by 10(-5) M nicotine was inhibited 70-80% by SP, SP-methyl ester, and the C-terminus-extended analogue SP-Tyr12-NH2, 65% by (Ala3)SP-NH2, 45% by the C-terminal analogue SP(4-11), and 20 and 5% by the N-terminal analogues SP(1-7) and SP(1-5), respectively, when these peptides were present at 3 x 10(-5) M concentrations. The order of potency was SP = SP-methyl ester = SP-Tyr12-NH2 > (Ala3)SP-NH2 > SP(4-11) > SP(1-7) > SP(1-5). SP, SP-methyl ester, and (Ala3)SP-NH2 protected against nicotinic desensitization by 40-55%, and SP(4-11) protected by 20% (all at 3 x 10(-5) M). In contrast, the N-terminal analogues SP(1-7) and SP(1-5) and the C-terminus-extended analogue SP-Tyr12-NH2 at 3 x 10(-5) M did not protect against nicotinic desensitization. Cyclo-SP(3-9), Ac-SP(3-9)-NH2, SP(3-9), and SP(3-6) had neither inhibitory nor facilitatory effects on secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

[Amalgams--discussion of a controversy]. / Amalgam--kontrovers diskutiert.

Amalgam fillings are still widely used in dentistry. Based on toxicological data, patient's and medical staff's risk of mercury poisoning is evaluated; pharmaco-prophylaxis against chronic poisoning is discussed.

Does a relationship exist between the quality of stress and the motivation to ingest alcohol?

Previous experimental findings on the relationship between emotional stress and motivation to ingest alcohol are contradictory. To obtain information about this relationship we tested the effects on alcohol consumption in rats subjected to two types of chronic unavoidable stressors, intermittent immobilization and social isolation, which differ in their influence on the functional state of the endogenous opioid system. To characterize the nature and magnitude of the stress induced by these stressors, we measured their effects on functional parameters which have a close relationship to the regulatory influence of endogenous opioid peptides (endogenous opioid dependence, pain sensitivity, blood pressure). Our investigations have shown that chronic intermittent immobilization, which induced development of endogenous opioid dependence, presumably due to activation of endogenous opioid systems, did not produce increased alcohol consumption. On the contrary, chronic social isolation, which did not induce development of endogenous opioid dependence, was followed by a significant increase in alcohol consumption. It is concluded that not all types of stress produce increased alcohol consumption, but that the effect on the endogenous opioid system may be a decisive factor in determining whether a stressor produces increased alcohol consumption.

In vivo-effect of intraadrenal nicotine and substance P application on rat adrenal medullary catecholamine secretion.

The present study was conducted to characterize in vivo the intraadrenal catecholamine (CA) secretion in rats. This was possible by using a microdialysis system (MDS) which mimics some properties of an artificial capillary. One end of this system was connected to a peristaltic pump, from the other end fractions were sampled at 5 min intervals. Concentrations of epinephrine (E) and norepinephrine (NE) in adrenal dialysate fractions were determined by HPLC electrochemical detection. Through this MDS nicotine was administered directly into the adrenal medulla of freely moving rats and the response of catecholamine release was determined. In the second part of the study the effect of exogenous substance P (SP) on spontaneous as well as on nicotine-stimulated CA release was investigated. Like nicotine, SP was administered directly into the adrenal medulla. At a flow rate of 25 microliter/min the transfer rates of CA and nicotine were approximately 1% whereas SP passed at a rate of 01.-0.2%. Under resting conditions CA release remained constant. In response to 2 x 10(-7) M nicotine (which resulted in local concentration of 2 x 10(-7) M), E and NE secretion increased 2.9 and 5.4-fold, respectively. However, due to an increased E response this difference attenuated with a later onset of the first stimulus. The higher concentrations of 10(-4) M resulted in 8.1 and 10.8-fold increases for E and NE. This latter response is clearly supraphysiologic and therefore the 2 x 10(-5) M concentration was used for further experimentation. CA secretion was stimulated with nicotine four times at 30 min intervals.(ABSTRACT TRUNCATED AT 250 WORDS)

Substance P and beta-endorphin-like immunoreactivity in lavage fluids of subjects with and without allergic asthma.

Six atopic subjects with grass pollen allergy and six nonallergic healthy volunteers were enrolled into this study. Substance P-like immunoreactivity (SP-LIR) and beta-endorphin-like immunoreactivity (beta E-LIR) were determined in bronchoalveolar lavage (BAL) and nasal lavage (NAL) fluids before and after allergen (grass pollen) provocation. A significant increase in the baseline concentration of SP-LIR and beta E-LIR was seen in BAL of allergic subjects. In NAL of allergic subjects an increased baseline concentration of SP-LIR was found (beta E-LIR not detectable). After allergen provocation there was a rise of SP-LIR and beta E-LIR in BAL fluids of allergic subjects immediately after provocation. In NAL fluids of allergic subjects allergen challenge resulted in a rise of SP-LIR within 10 minutes. Allergen provocation did not influence SP-LIR and beta E-LIR concentration in BAL and NAL in nonallergic controls. The demonstrated higher baseline levels of SP-LIR and beta E-LIR as well as the increase after provocation in the BAL and NAL of allergic subjects but not in nonallergic controls support the hypothesis that these neuropeptides contribute to allergic reactions in airways of humans.

Endogenous opioid dependence--a basic pathophysiological phenomenon of stress-induced and genetically fixed disturbances in adaptation--influence of substance P.

Disturbances in adaptive processes can be induced by chronic exposition to stress or can result from a genetical predisposition. Experimental data of chronically stressed Wistar rats and of spontaneously hypertensive rats (SHR) demonstrate a relation between a decreased level of substance P (SP) in adrenals, the existence of a dependence on endogenous opioid peptides and an increased regulatory level of blood pressure. The endogenous level of SP was determined by using a RIA. The dependence (physical) on endogenous opioid peptides was detected by using the method of "gut dependence". SP injection i.p. once a day for 4 d antagonized the dependence on endogenous opioid peptides and normalized the increased level of blood pressure in both animal models. Investigations on SHR had shown that the adaptive effect of SP on blood pressure and endogenous opioid dependence is bound to the premise of an acute stimulated endogenous opioid system at the moment of SP-application. Experimental findings suggest that different systems of opioid peptides take part in the etiopathogenesis of genetically predisposed hypertension of SHR and in stress-induced increase of blood pressure level of Wistar rats. The effect of SP on blood pressure and endogenous opioid dependence will be discussed as a result of the modulatory influence on the cholinergic-opioid-peptidergic interaction.

[The delta sleep-inducing peptide as a factor enhancing the content of substance P in the hypothalamus and the resistance of rats to emotional stress]. / Peptid, vyzyvaiushchii del'ta-son, kak faktor, povyshaiushchii soderzhanie veshchestva P v gipotalamuse i ustoichivost' krys k émotsional'nomu stressu.

The influence of the delta-sleep inducing peptide (DSIP, 60 and 120 nmol/kg, intraperitoneally) on the content of substance P (SP) in rats hypothalamus was studied on males of August line. DSIP administration significantly increased the mean SP content in the hypothalamus and also its content in animals, stable and predisposed to emotional stress. Daily DSIP administration before putting the rats in conditions of stress increased the SP content in the hypothalamus decreased at the emotional stress. Preliminary single DSIP administration to the animals subjected to stress also increased the SP content. Single DSIP administration in a dose of 60 nmol/kg sharply reduced classical stress manifestations, such as hypertrophy of adrenals and thymus involution.

The possible role of substance P in the allergic reaction, based on two different provocation models.

It was shown in two different provocation models (nasal and bronchial provocation) that substance P (SP) may play an important role in the neurogenic inflammatory response in upper and lower airway disease. (1) Pretreatment with SP augments the antigen challenge response of the nasal mucosa. (2) The baseline bronchoalveolar lavage (BAL) concentrations of SP are elevated 8-fold in allergies (pollen asthma) as compared with normals, even outside of season. (3) The SP concentration in BAL increases significantly (p less than 0.05) after bronchial allergen provocation. These findings support a previous hypothesis of an abnormally elevated activity of nonadrenergic-noncholinergic excitatory nerves and are in accordance with the results of a decreased activity of neutral endopeptidase exaggerating neurogenic inflammatory responses in the airways, including bronchomotor tone hyperresponsiveness.

[Beta-endorphin and substance P in the perioperative period]. / Beta-Endorphin und Substanz P in der perioperativen Periode.

The modifying impact of anaesthesia on the stress reaction related to surgical trauma was investigated on the basis of the neuropeptidergic parameters of 66 patients who had to undergo a gynaecological radical operation. Anaesthesia was either performed as neuroleptanaesthesia or as epidural analgesia by using bupivacaine in combination with general anaesthesia. The plasma concentrations of substance P and beta-endorphin were taken as neuropeptidergic parameters. Both regulatory peptides show numerous corresponding synergisms. An acceleration of these neuropeptide systems is assumed to be present in severe disturbance of homeostasis. Plasma concentrations of substance P and beta-endorphin were examined at 11 measuring points in the perioperative and intraoperative periods. The plasma concentration of substance P significantly declines in the preoperative period while the concentration of beta-endorphin in the plasma remains at a relatively constant level. In the dynamics of beta-endorphin in the plasma significant differences between the two anaesthetic techniques become apparent in the intraoperative period. Those patients given epidural analgesia have a significantly higher maximum concentration at a later date. This difference is attributed to the possible loss of the adrenal medullary function due to partial sympathetic blocking. Single observations in patients pregnant in the last trimester testify to an extraordinary adaptability at the end of pregnancy.

Stress-induced dependence on endogenous opioid peptides--a fundamental process in the pathophysiology of a disturbed adaptation--its influence by substance P.

Using previous findings of stress-induced disturbances in regulation of peripheral and central function within the general adaptation process and hints from the literature to a multiple participation of opioid peptides in adaptive processes, this paper represents experimental data which gives hints to a functional relation in the etiopathophysiology between the development of stress-induced dependence on endogenous opioid peptides and stress-induced disturbances in blood pressure regulation and to the adaptive effect of substance P within the peptidergic interaction with endogenous opioid peptides and its importance for the physiology or pathophysiology of adaptive processes.

Possible role of substance P on regulation of pituitary-adrenal axis.

The effects of chronic immobilization stress on the physiological responses of male rats were studied. The results indicate that the SP-like immunoreactivity (SPLIR) is diminished in the adrenals and pituitary after chronic stress. In vitro noradrenaline (NA) release from adrenals was increased. The i.p. administration of SP during the stress procedure normalized the increased NA release in vitro indicating that the catecholamine secretion may be influenced by SP. On the other hand, in demedullated animals the SPLIR in the pituitary was partly reduced and the blood pressure was increased. In such animals chronic stress resulted in an increase of SPLIR in the pituitary in comparison with nonstressed, demedullated animals, but was without effect on the blood pressure. It is concluded that exposure to SP and the resulting decrease of noradrenaline release may have a significant influence on the pituitary-adrenal responsiveness to stress.

Postnatal development of substance P in the inner ear of the guinea pig.

Appreciable amounts of substance P (SP) were found in guinea pig cochleas. The highest values were found in the postnatal period. Data presented favor the assumption of SP acting as a neuromodulator or neurotransmitter in the inner ear.