Cell relationship in a Wistar rat model of spontaneous prostatitis.

PURPOSE: Prostatitis in men is a painful, noninfectious inflammatory condition. It is similar to interstitial cystitis which is associated with increased bladder mast cell and sensory nerve fiber density as well as suprapubic pain. Certain strains of rats may provide a useful model for studies of the development of spontaneous prostatitis. We evaluated the time course, and involvement of mast cells and sensory nerve fibers in this process using Wistar rats. MATERIALS AND METHODS: The prostates of 4, 6, 8, 10 and 13-week-old male Wistar rats were examined for the degree of inflammation, innervation, mast cell density and nerve mast cell relationship using histochemical and immunocytochemical studies. Bacterial cultures of tissue were performed at 13 weeks. RESULTS: The inflammatory cell index increased progressively with age. Inflammation was moderate and consisted mostly of lymphocytes and macrophages associated with occasional glandular epithelial necrosis and edema. The density of nerve fibers immunoreacting with the neuronal marker protein gene produce 9.5 increased gradually with age and fibers immuno-positive for the sensory neuropeptide calcitonin gene-related peptide more than doubled by 13 weeks compared with by 4 weeks. The density of visible mast cells declined after 4 weeks in a pattern that corresponded with the increased percent of mast cells undergoing degranulation. For the mast cells with calcitonin gene-related peptide immuno-positive nerve fibers within a distance of 40 microm. distance correlated significantly with the degree of degranulation. Bacterial cultures were negative at 13 weeks. CONCLUSIONS: Our results confirm previous reports of spontaneous prostatitis in Wistar rats and indicate that moderate inflammation may occur in 80% of rats at as early as age 13 weeks. While the correlation of the nerve mast cell axis with mast cell degranulation does not prove our hypothesis of mast cell mediated inflammatory mediator release in the development of nonbacterial prostatitis, it suggests that such a relationship is possible.

mRNA expression of novel CGRP1 receptors and their activity-modifying proteins in hypoxic rat lung.

Calcitonin gene-related peptide (CGRP) is a potent vasodilator. Our group has reported that exogenous CGRP may prevent or reverse hypoxic pulmonary hypertension in rats. The vasodilatory action of CGRP is mediated primarily by CGRP1 receptors. The calcitonin receptor-like receptor (CRLR) and the orphan receptor RDC-1 have been proposed as CGRP1 receptors, and recent evidence suggests that CRLR can function as either a CGRP1 receptor or an adrenomedullin (ADM) receptor. Receptor activity-modifying proteins (RAMPs) determine the ligand specificity of CRLR: coexpression of CRLR and RAMP1 results in a CGRP1 receptor, whereas coexpression of CRLR and RAMP2 or -3 results in an ADM receptor. We used qualitative, semiquantitative, and real-time quantitative RT-PCR to detect and quantitate the relative expression of these agents in the lungs of rats exposed to normoxia (n = 3) and 1 and 2 wk of chronic hypobaric hypoxia (barometric pressure 380 mmHg, equivalent to an inspired O(2) level of 10%; n = 3/time period). Our results show upregulation of RDC-1, RAMP1, and RAMP3 mRNAs in hypoxic rat lung and no change in CRLR and RAMP2 mRNAs. These findings support a functional role for CGRP and ADM receptors in regulating the adult pulmonary circulation.

Three-week neonatal hypoxia reduces blood CGRP and causes persistent pulmonary hypertension in rats.

To increase understanding of persistent pulmonary hypertension, we examined chronic pulmonary effects of hypoxia at birth and their relationships with immunoreactive levels of the potent vasodilator, calcitonin gene-related peptide (CGRP). Rats were born in 10% hypobaric hypoxia, where they remained for 1-2 days, or in 15% hypoxia, where they remained for 21 days. All were then reared in normoxia for 3 mo followed by reexposure to 10% hypoxia for 7 days (H-->H) or continued normoxia (H-->N); age-matched normoxic rats were hypoxic for the last 7 days (N-->H) or normoxic throughout (N-->N). Results are as follows. Pulmonary arterial pressure (P(PA)) in 10% H-->N rats was normal at the end of the experiment (13 wk), but in rats reexposed to hypoxia (H-->H), pressure rose to 19% above N-->H controls. In 15% H-->N rats, P(PA) remained high, similar to that of N-->H rats, and increased further by 40% on reexposure (H-->H). Medial thickness of small pulmonary arteries in 10% H-->H rats also increased by 40% over N-->H controls and was equally high in 15% H-->N and H-->H rats. In N-->H rats from both experiments, right ventricular hypertrophy index (RVH) was increased after hypoxia at 15-16 wk. Also, in the 15% study, RVH remained elevated in H-->N rats and increased in H-->H rats by 19% above N-->H controls. Blood CGRP was reduced by neonate and adult hypoxia, and hypoxic reexposure (H-->H) further lowered blood CGRP in the 15% but not 10% study. Declining left ventricular blood CGRP correlated highly with logarithmically increasing P(PA) in the 15% study (r = -0.81, P = 0.000). In conclusion, 1) short perinatal exposure to 10% O(2) exacerbated pulmonary hypertension with hypoxia later in life, 2) 15% O(2) at birth and for 21 days caused persistent pulmonary hypertension and exacerbation with reexposure, and 3) P(PA) correlated highly with declining blood CGRP levels in the 15% study.

5-HT5a receptors in the carotid body chemoreception pathway of rat.

By using a specific antibody, 5-HT5a receptor-like immunoreactivity was revealed in the chemoreceptive, oxygen sensitive, carotid body (CB) type I cells, and neurons of the petrosal ganglion (PG) and the superior cervical ganglion (SCG) in rat. mRNA encoding for the 5-HTa receptor was also detected in these tissues by RT-PCR, and confirmed with DNA sequencing. The present study provides direct evidence that 5-HT5a receptors are expressed in the CB, PG and SCG, which all likely play fundamental roles in arterial chemoreception.

The role of endogenous lung neuropeptides in regulation of the pulmonary circulation.

Vascular resistance in the mammalian pulmonary circulation is affected by many endogenous agents that influence vascular smooth muscle, right ventricular myocardium, endothelial function, collagen and elastin deposition, and fluid balance. When the balance of these agents is disturbed, e.g. by airway hypoxia from high altitude or pulmonary obstructive disorders, pulmonary hypertension ensues, as characterized by elevated pulmonary artery pressure (P(PA)). Among neuropeptides with local pulmonary artery pressor effects are endothelin-1 (ET-1), angiotensin II (AII), and substance P, and among mitigating peptides are calcitonin gene-related peptide (CGRP), adrenomedullin (ADM), atrial natriuretic peptide (ANP), vasoactive intestinal peptide (VIP) and ET-3. Moreover, somatostatin28 (SOM28) exacerbates, whereas SOM14 decreases P(PA) in hypoxic rats, with lowering and increasing of lung CGRP levels, respectively. Pressure can also be modulated by increasing or decreasing plasma volume (VIP and ANP, respectively), or by induction or suppression of vascular tissue remodeling (ET-1 and CGRP, respectively). Peptide bioavailability and potency can be regulated through hypoxic up- and down- regulation of synthesis or release, activation by converting enzymes (ACE for AII and ECE for ET-1), inactivation by neutral endopeptidase and proteases, or by interaction with nitric oxide (NO). Moreover, altered receptor density and affinity can account for changed peptide efficacy. For example, upregulation of ET(A) receptors and ET-1 synthesis occurs in the hypoxic lung concomitantly with reduced CGRP release. Also, receptor activity modifying protein 2 (RAMP2) has been shown to confer ADM affinity to the pulmonary calcitonin-receptor-like receptor (CRLR). We recently detected the mRNA encoding for RAMP2, CRLR, and the CGRP receptor RDC-1 in rat lung. The search for an effective, lung selective treatment of pulmonary hypertension will likely benefit from exploring the imbalance and restoring the balance between these native modulators of intrapulmonary pressure. For example, blocking of the ET-1 receptor ET(A) and vasodilation by supplemental CGRP delivered i. v. or via airway gene transfer, have proven to be useful experimentally.

Sensory CGRP depletion by capsaicin exacerbates hypoxia-induced pulmonary hypertension in rats.

Pulmonary hypertension is a debilitating disease that occurs among infants and adults. One of many etiologies is airway hypoxia. We previously demonstrated a role of endogenous calcitonin gene-related peptide (CGRP), a potent vasodilator, in ameliorating the pulmonary vascular pressor response to chronic hypoxia and related changes in the lungs and heart. This study evaluates the role of endogenous sensory CGRP in hypoxic pulmonary hypertension and examines the intrinsic neural microcircuitry. Rats were pretreated with capsaicin i.p. to deplete pulmonary sensory C-fiber stores of CGRP and substance P and placed in hypobaric hypoxia (10% O2, 16 days) or normoxia together with sham controls. Hypoxia increased pulmonary artery pressure, right-ventricular weight, arterial medial thickness, elasticized capillaries, endothelial cell density, lung water and hematocrit in control rats. Capsaicin augmented pulmonary artery pressure and right-ventricular hypertrophy in hypoxia, and medial thickness and endothelial cell density both in normoxia and hypoxia. Because of the limited effects on these parameters by substance P and other capsaicin-sensitive lung agents, our results suggest that sensory CGRP deficit severely exacerbates pathological signs of hypoxic pulmonary hypertension. A neural microcircuitry consistent with an axon reflex pathway is outlined histochemically. We conclude that endogenous CGRP modulates pulmonary vascular tone in hypoxic pulmonary hypertension which requires intact primary sensory fibers.

A1-adenosine receptor antagonists block endotoxin-induced lung injury.

Endotoxin produces a variety of biological effects on different cell types, such as priming of neutrophils and macrophages, which then release a number of important mediators of endotoxin-induced lung injury. However, the specific mechanism by which endotoxin initiates its cascade of pathophysiological events in the lung has not been described. Both A1 adenosine receptor activation and endotoxin induce the release of thromboxane A2 from the lung and inhibit adenylate cyclase. By acting on A1 adenosine receptors, adenosine promotes neutrophil chemotaxis and adherence to endothelial cells. We hypothesized that A1 adenosine receptor activation is essential to endotoxin-induced lung injury, and we used the highly selective A1-adenosine receptor antagonists, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) and 8-benzyl-7,[2-[ethyl(2-hydroxyethyl)amino]-ethyl] theophylline (bamiphylline), to investigate whether selective blocking of the A1 adenosine receptor would prevent endotoxin-induced acute lung injury. An intralobar arterial infusion of endotoxin (15 mg/kg) into the left lower lobe of the lung in intact-chest, spontaneously breathing cats produced lung injury characterized by the presence of neutrophils, macrophages, and red blood cells (RBCs) in alveoli, and alveolar edema and necrosis. Lower doses of endotoxin (5 or 10 mg/kg) produced less severe and dose-dependent lung injury. Endotoxin (15 mg/kg)-induced alveolar injury was blocked in a highly significant manner by A1-adenosine receptor antagonists, DPCPX and bamiphylline. An intravenous bolus of DPCPX 30 min before endotoxin infusion or a continuous intravenous infusion of bamiphylline 30 min before, during, and 30 min after endotoxin reduced the percent injured alveoli (defined as the presence of 2 or more inflammatory cells or RBCs, or edematous fluid) from 57 +/- 31% (endotoxin 15 mg/kg) to 9 +/- 1% (DPCPX) or 21 +/- 14% (bamiphylline), which were not significantly different from control (1-h perfusion only) (4 +/- 1%) (P < 0.05). These data represent the first evidence that A1-adenosine receptor antagonism blocks the capacity of endotoxin to cause lung injury. A1-adenosine receptor antagonists may be useful in preventing adult respiratory distress syndrome associated with septicemia.

Intramyocardial arterial cushions of coronary vessels in animals and humans: morphology, occurrence and relation to heart disease.

The existence of coronary endoarterial cushions (CEC) in the human heart as nonpathological, functional entities has been debated, and CEC have been sparsely reported in animals. Arterial cushions are localized thickenings that protrude into the lumen of specific arteries. We have identified CEC in the rhesus monkey, dog, sheep, goat, pig, rabbit and rat, and in the human heart. Two distinct types are described: the ovoid CEC arranged singly, in pairs, or in groups of three to four, and the less common polypoid CEC seen primarily in humans. The highest incidence of CEC in rabbits and humans was in the left ventricle in arteries 150-488 microns in diameter. Light and electron microscopy demonstrated intimal location with smooth muscle cells surrounded by ground substance, collagen and elastin fibers in a highly organized pattern. Nerve fibers identified by their immunoreactivity with antiserum to the vasodilatory calcitonin-gene-related peptide contacted the CEC along the tunica media and were occasionally seen within CEC. Arrangement and histological composition of CEC suggest a role in the regulation of local blood flow and myocardial perfusion. In human hearts, the CEC density index correlated highly with the degree of heart disease. In subjects with high heart disease rating, increased connective tissue, lipid-like infiltration and calcification was seen within CEC, and foam cells were present in CEC of obese rabbits. This suggests that CEC in coronary arteries could be predisposed sites of atherosclerosis, and that injured CEC can cause coronary artery spasm and ischemia. We conclude that CEC occur in animals and humans as innervated intimal smooth muscle cushions that might have a role in myocardial perfusion and heart disease.

Nerve-mast cell interaction in normal guinea pig urinary bladder.

Urinary bladders of normal adult female guinea pigs were analyzed for anatomical evidence of nerve-mast cell interaction using light microscopy and electron microscopy. Nerves, ganglia, and individual nerve fibers were visualized on paraffin sections using immunohistochemistry with antisera against the neural antigens neurofilament protein and protein gene product 9.5, and sections were also immunoreacted with antisera against the neuropeptides substance P and calcitonin gene-related peptide. Separate mast cell populations were identified by counterstaining with toluidine blue and alcian blue. Mast cells of both types were found within nerves and intramural ganglia and were in close contact with individual nerve fibers displaying substance P- and calcitonin gene-related peptide-like immunoreactivity. Moreover, serotonin-immunoreactive mast cells were innervated with nerve fibers that reacted with antiserum against vasoactive intestinal polypeptide. At the ultrastructural level, these fibers were almost exclusively identified as unmyelinated primary sensory afferents. Mast cells contacted these fibers with lamellipodia that wrapped around and enclosed the fibers deeply within the cell. Close association between mast cells, nerves, and vessels was common. Ultrastructural evidence suggests that bidirectional communication occurs between nerve fibers and mast cells. These structures may participate in axon reflexes that regulate normal vascular and detrusor smooth muscle function and cause vasodilation, edema, inflammation, and bladder hyperreactivity. In summary, a close relationship exists between mast cells and peptidergic nerve fibers, including primary sensory afferents. Results suggest that bidirectional interaction could occur between nerves and mast cells.

Intracarotid substance P infusion inhibits ventilation in the goat.

Substance P (SP) has been proposed as an excitatory neuromodulator of the carotid body (CB) response to hypoxia based on data from the cat and rat. The role of SP as a CB neuromodulator in the goat is unknown. Awake (n = 14) and chloralose anesthetized goats (n = 6) were used to investigate the effects of intracarotid (IC) SP infusions (1-6 micrograms.kg-1.min-1) and bolus injections (6 micrograms kg-1) to the CB intact and denervated (CBX) sides (control) on mean ventilation (VE) and mean blood pressure (MBP). In awake goats VE was decreased by infusion or bolus SP injection at a dose of 6 micrograms.kg-1 (P < 0.05) and occurred with infusions to the intact or CBX sides. MBP was elevated with SP infusion to either the CB intact or CBX sides at all SP doses. The SP antagonist CP-96,345 (0.1 mg.kg-1, IV) blocked the decrease in VE induced by SP in normoxia and significantly increased the hypoxic ventilatory response (PaO2 = 40 torr). In anesthetized goats, IC injections of SP (1 to 6 micrograms.kg-1) reduced phrenic activity and MBP before and after CBX. In only one of five goats airway pressure was increased suggesting that bronchoconstriction was not a cause for the reduced ventilatory and phrenic activity induced by SP. Immunohistochemistry provided evidence of SP in CB nerve fibers and terminals, carotid sinus nerve axons and petrosal ganglion cells, but not in type I glomus cells. Our results do not support the view that SP is an excitatory neuromodulator of CB chemotransduction in the goat.

A1 adenosine receptor antagonists block ischemia-reperfusion injury of the lung.

Ischemia-reperfusion (I-R) injury of the lung occurs after lung transplantation, pulmonary thromboembolectomy, or cardiopulmonary bypass. In the heart, adenosine, A1 adenosine receptor agonists, and a brief period of preconditioning ischemia attenuate I-R injury. Moreover, in the lung, thromboxane is released during ischemia and is an important mediator of I-R injury. We previously reported that adenosine produces vasoconstriction in the feline pulmonary vascular bed by acting on A1 receptors to induce the release of thromboxane and that these vasoconstrictor responses are desensitized by low doses of A1 receptor agonists. Because A1 receptor agonists mimic the effect of preconditioning ischemia, we hypothesized, in contrast to previously proposed mechanisms, that small amounts of adenosine released during preconditioning ischemia desensitize A1 receptors. Also, we hypothesized that greater amounts of adenosine are released after longer periods of ischemia, which activate A1 receptors. Thus if desensitization of A1 receptors is the mechanism by which preconditioning attenuates I-R injury of the heart and A1 receptor activation during ischemia plays an important role in I-R injury of the lung, A1 receptor antagonists should provide a protective effect in I-R injury of the lung. In this study, 2 h of ischemia and 2 h of reperfusion of the left lower lobe in intact-chest, spontaneously breathing cats caused lung injury characterized by the presence of neutrophils, macrophages, and RBCs in alveoli and caused alveolar edema, which was blocked in a highly significant manner by the A1 receptor antagonists xanthine amine congener (XAC) and 1,3-dipropyl-8-cyclopentylxanthine (DPCPX). An intralobar arterial infusion of XAC (30 min before ischemia) reduced the %injured alveoli (defined as presence of 2 or more inflammatory cells or RBCs, or edematous fluid) from 60 +/- 10 to 7 +/- 2%, which was not significantly different from controls (5 +/- 1%; P < 0.0001). DPCPX (iv) reduced the %injured alveoli to 13 +/- 7% when administered 30 min before ischemia and to 6 +/- 2% when administered after 1 h of reperfusion, not significantly different from controls (P < 0.0001). Preconditioning ischemia (10-min ischemia +10-min reperfusion) also reduced the %injured alveoli after 2 h ischemia and 2 h reperfusion to 23 +/- 13%, almost identical to 2 h ischemia and 1 h reperfusion. These data support the hypothesis that A1 receptor antagonists block I-R injury of the lung. A1 receptor antagonists may be useful in preventing I-R injury after transplant surgery and during surgical procedures associated with I-R injury of the heart, brain, kidney, and spinal cord.

Differential release of prostaglandins and leukotrienes by sensitized guinea pig urinary bladder layers upon antigen challenge.

The relative contributions of mucosal/submucosal and detrusor layers to the release of inflammatory mediators were investigated in isolated, ovalbumin (OVA) sensitized guinea pig urinary bladders. Ovalbumin challenge of sensitized mucosa induced release of prostaglandins (PG): PGE2, PGD2 and PGF2 alpha, in that order of magnitude. The total release of PGs was significantly higher from the mucosa/submucosa than from the detrusor/serosa. Under the same conditions, net release of leukotriene (LT) was observed predominantly from the detrusor. The total amount of histamine released from the mucosa was greater than that from the muscle layer. These results indicate differential production and release of inflammatory mediators from the mucosal/submucosal and detrusor smooth muscle layers. These results may have serious implications in disorders, such as interstitial cystitis, involving bladder mucosal damage. The cytoprotective effect of PGE2 is likely to be lost when the mucosa is damaged, and LT release from deeper layers may contribute significantly to symptoms of bladder inflammation.

Reversal of cytochrome P-4501A1 and P-450-EF expression in MCA-C3H/10T1/2 cell-derived tumors as compared to cultured cells.

The 3-methylcholanthrene-transformed tumorigenic cell line, MCA-C3H/10T1/2 CL15 (MCA), expresses the novel benz(a)anthracene (BA)-inducible polycyclic aromatic hydrocarbon-metabolizing cytochrome P-450 (P-450-EF). The level of expression is comparable to that reported for the nontumorigenic C3H/10T1/2 CL8 (10T1/2) cells (Pottenger, L. H., Christou, M., and Jefcoate, C. R. Arch. Biochem. Biophys., 286: 488-497, 1991). Sarcomas (3-12 mm in diameter) generated in athymic "nude" mice by s.c. injection of MCA cells exhibited much lower 7,12-dimethylbenz(a)anthracene-metabolizing activities (5-15% of the levels in cultured cells), both constitutively and after in vivo treatment with BA. A sharp decrease in P-450-EF expression was observed both at the functional level (10- to 30-fold) (as determined by antibody inhibition studies) and at the apoprotein level (50- to > 100-fold) (as determined by Western immunoblots). However, in contrast to the BA-treated MCA cells in which P-450-EF comprises essentially the total spectrally detectable P-450 content (approximately 30 pmol/mg) with virtually undetectable cytochrome P-4501A1, tumors from these cells expressed substantial levels of P-4501A1 immunodetectable protein in response to BA treatment (approximately 0.5-3 pmol/mg). In these tumors, P-450-EF expression decreased to undetectable or barely detectable levels (< 0.2-0.5 pmol/mg). P-4501A1-expressing cells were localized in tumor sections immunocytochemically and were morphologically identical to other MCA cells, which formed the majority of the sarcoma. At the functional level, antibody inhibition studies and product ratios demonstrated that P-4501A1 accounted for only 40% of the total dimethylbenz(a)anthracene-metabolizing activity of BA-induced tumor microsomes, whereas the remaining activity was due to P-450-EF. This low catalytic activity for P-4501A1 (35-95 pmol/mg/h) indicated that the majority of BA-inducible P-4501A1 in the tumors (> 95%) was expressed as apoprotein. The contribution from P-450-EF was consistent with full expression of hemoprotein. Tumor size affected the total dimethylbenz(a)anthracene-metabolizing activities/mg microsomal protein (small tumors were 2- to 3-fold more active than large tumors) but had no effect on the ratio of activities dependent on, respectively, P-450-EF and P-4501A1 holoenzymes (1.5:1), thus suggesting controlled coexpression of these proteins. Reculturing of tumor-derived cells effected partial restoration of P-450-EF expression and eliminated the expression of P-4501A1, confirming a unique contribution from tumor environment to the regulation of these genes. Possible mechanisms for an environment-dependent concerted regulation of the P-4501A1 and P-450-EF genes are discussed.

In vitro effects of bladder mucosa and an enkephalinase inhibitor on tachykinin induced contractility of the dog bladder.

Tachykinin-induced contractility of smooth muscle strips from dog bladders was studied in vitro, and the presence of substance P-like immunoreactivity and neurokinin A and neurokinin B-like immunoreactivity was examined in bladder sections. Nerve fibers with substance P-like immunoreactivity were present in the mucosa, submucosa and smooth muscle. Fibers were also found in nerves, intramural ganglia, and around blood vessels. Neurokinin A-like immunoreactivity had similar distribution, and no neurokinin B-like immunoreactivity was observed. Removal of the mucosa significantly enhanced the sensitivity and the maximum responses to the tachykinins. After removing the mucosa, the sensitivity to these tachykinins increased 0.4 to 0.5 log units (p less than 0.02). The responses to carbachol were not altered by mucosa removal. The leftward shifts of the concentration-response curves for neurokinin A were of similar magnitude after removal of the mucosa, and after pretreatment with phosphoramidon (10 microM), an enkephalinase inhibitor, in the presence of mucosa. However, phosphoramidon did not alter the sensitivity of the bladder strips to neurokinin B, and slightly changed the sensitivity to substance P (0.2 log units). Additional shifts of the substance P and neurokinin A curves to the left were observed in the presence of phosphoramidon when the mucosa was removed (0.6 and 0.5 log units, p less than 0.005). The order of potency for the tachykinins (neurokinin A greater than substance P) was not altered by mucosa removal, addition of phosphoramidon, or both. Neurokinin A was degraded by enkephalinase located in the bladder mucosa and addition of phosphoramidon or mucosa removal resulted in an inhibition or loss of enkephalinase activity. It is concluded that the responses to neurokinin A, which acts on NK-2 type of receptors, prevail on the dog bladder.

Dynamic aspects of regulatory lung peptides in chronic hypoxic pulmonary hypertension.

Male Sprague-Dawley rats were placed in hypobaric hypoxia for 17-21 d (FIO2 10%) to establish pulmonary hypertension (PH) and control rats were kept in normobaric room air. Right mean atrial and ventricular pressures (PRA, PRV) were recorded, left ventricular (LV) blood was collected, and lungs were perfused with heparinized saline. Hearts were removed to evaluate right ventricular (RV) hypertrophy (RV/(LV+septum)%). Peptides were quantitated with radioimmunoassay in lung tissue extracts and plasma. Wet lung weight, PRA, PRV, and RV/(LV+S)% were higher and body weight was lower in hypoxia rats, and lung morphometry revealed increased arterial medial thickness (MT/OD%) and elastification of arterioles and capillaries. Lung tissue CGRP, PYY, gamma 2-MSH, and SOM were higher in PH rats and ANP was unchanged. Blood AVP, CGRP, PYY, VIP, and SOM were reduced in PH rats and ANP was unchanged. Lung levels of PYY and SOM correlated significantly with the time in hypoxia and with all parameters examined and CGRP and gamma 2-MSH correlated with all but medial thickness. PYY had the highest correlation of the peptides with body weight, PRV, and RV/(LV+S)%, and SOM the highest with time in hypoxia, wet lung weight, PRA, MT/OD%, and elastification of arterioles and capillaries. Blood peptides correlated inversely with these parameters. ANP had the overall weakest correlations and CGRP, PYY, and SOM had the highest. SOM correlated the highest with arterial medial hypertrophy, PRV, RV hypertrophy, and elastification of peripheral capillaries. VIP correlated the highest of the blood peptides with body weight and wet lung weight. Statistically significant correlations do not necessarily imply causal relationships. The putative roles of these peptides in pulmonary function are discussed.

Calcitonin gene-related peptide and its mRNA in pulmonary neuroendocrine cells and ganglia.

The occurrence of calcitonin gene-related peptide (CGRP) and it's mRNA was studied in lungs of rats and piglets using in situ hybridization with two synthetic oligonucleotide probes followed by immunocytochemistry (ICC). CGRP mRNA was present in pulmonary neuroendocrine cells (PNEC) of both the solitary type and cluster type (neuroepithelial body; NEB) at all levels of the airway epithelium from bronchi to alveoli. The distribution of labelled cells was similar to that previously described with ICC. The 44-mer probe provided stronger hybridization signal than the 34-mer and the two combined increased labelling slightly. Formalin fixation reduced labelling and tended to increase background. Labelling for CGRP mRNA was evenly distributed over the cytoplasm, whereas CGRP-like immunoreactivity (LI) usually was of highest intensity toward the base of the PNEC, suggesting basal accumulation of synthesized peptide. CGRP-LI was also observed in occasional rat ganglia and in some, but not all, piglet ganglia. These local neurons may contribute to the CGRP fibers of airways and vasculature, and could theoretically bridge their dendrites and axons between NEB and the effector organ (e.g. artery or arteriole) thus accomplishing a function similar to the postulated axon reflex.

PYY-like material and its spatial relationship with NPY, CGRP and 5-HT in the lung of the Syrian golden hamster.

We investigated the presence of peptide YY, neuropeptide Y, calcitonin gene-related peptide and serotonin in the hamster lung by radioimmunoassay, high performance liquid chromatography and immunocytochemistry. Lung-tissue concentrations of peptide YY and neuropeptide Y were 1.3 +/- 0.2 and 2.5 +/- 0.2 pmol/g wet weight, respectively. These two closely related pancreatic peptides were demonstrated in separate peaks with high performance liquid chromatography. The peptide YY appeared fragmented as immunoreactive peptide YY eluted primarily late in the gradient but showed additional peaks early in the gradient. Peptide YY-like immunoreactivity (PYY-LI) was predominantly observed in one or more cells of neuroepithelial bodies in all airways peripheral to bronchioles, and in solitary neuroendocrine cells primarily located in the same peripheral areas. Neuropeptide Y-LI was seen in individual, thin nerve fibers around arteries and veins, in the airway lamina propria, and in the airway epithelium; in the latter also immunopositive nerve terminals were located. This pattern did not appear to coincide with that of calcitonin gene-related peptide-LI in epithelial nerve fibers and terminals. Peptide YY-LI, calcitonin gene-related-LI and serotonin-LI were present in cells of one and the same neuroepithelial body. However, peptide YY-LI was never found to be co-localized with calcitonin gene-related-LI or serotonin-LI, but the latter two were co-localized as previously reported.