Side-stream tobacco smoke-induced airway hyperresponsiveness in early postnatal period is involved nerve growth factor.

Epidemiological studies have shown that children are more susceptible to adverse respiratory effects of passive smoking than adults. The goal of this study is to elucidate the possible neural mechanism induced by exposure to passive smoking during early life. Postnatal day (PD) 2 and PD 21 mice were exposed to side-stream tobacco smoke (SS), a surrogate to secondhand smoke, or filtered air (FA) for 10 consecutive days. Pulmonary function, substance P (SP) airway innervation, neurotrophin gene expression in lung and nerve growth factor (NGF) release in bronchoalveolar lavage (BAL) fluid were measured at different times after the last SS or FA exposure. Exposure to SS significantly altered pulmonary function in PD2, accompanied with an enhanced SP innervation in airway. However, exposure to SS during the later developmental period (PD21) did not appear to affect pulmonary function and SP innervation of the airways. Interestingly, SS exposure in PD2 group significantly induced an increased gene expression on NGF, and decreased NGF receptor P75 in lung; parallel with high levels of NGF protein in BAL. Furthermore, pretreatment with NGF antibody significantly diminished SS-induced airway hyperresponsivenss and the increased SP airway innervation in the PD2 group. These findings suggest that enhanced NGF released in the lung contributes to SS-enhanced SP tracheal innervation and airway responsiveness in early life.

Early postnatal exposure of mice to side-steam tobacco smoke increases neuropeptide Y in lung.

Our recent study showed that prenatal and early postnatal exposure of mice to side-steam tobacco smoke (SS), a surrogate to environmental tobacco smoke (ETS), leads to increased airway responsiveness and sensory innervation later in life. However, the underlying mechanism initiated in early life that affects airway responses later in life remains undefined. The concomitant increase in nerve growth factor (NGF) after exposures suggests that NGF may be involved the regulation of airway innervation. Since NGF regulates sympathetic nerve responses, as well as sensory nerves, we extended previous studies by examining neuropeptide Y (NPY), a neuropeptide associated with sympathetic nerves. Different age groups of mice, postnatal day (PD) 2 and PD21, were exposed to either SS or filtered air (FA) for 10 consecutive days. The level of NPY protein in lung and the density of NPY nerve fibers in tracheal smooth muscle were significantly increased in the PD2-11SS exposure group compared with PD2-11FA exposure. At the same time, the level of NGF in lung tissue was significantly elevated in the PD2-11SS exposure groups. However, neither NPY (protein or nerves) nor NGF levels were significantly altered in PD21-30SS exposure group compared with the PD21-30FA exposure group. Furthermore, pretreatment with NGF antibody or K252a, which inhibits a key enzyme (tyrosine kinase) in the transduction pathway for NGF receptor binding, significantly diminished SS-enhanced NPY tracheal smooth muscle innervation and the increase in methacholine-induced airway resistance. These findings show that SS exposure in early life increases NPY tracheal innervation and alters pulmonary function and that these changes are mediated through the NGF.

A2A adenosine receptor deficiency leads to impaired tracheal relaxation via NADPH oxidase pathway in allergic mice.

A(2A) adenosine receptor (A(2A)AR) has been shown to suppress superoxide generation in leukocytes via the cAMP-protein kinase A (PKA) pathway. However, no study has yet explored the role of A(2A)AR in relation to NADPH oxidase in murine tracheas in vitro, which may lead to altered smooth muscle relaxation in asthma. Therefore, the present study evaluated the effects of A(2A)AR deficiency on the NADPH oxidase pathway in tracheas of A(2A) wild-type (WT) and A(2A) knockout (KO) mice. A(2A)WT mice were sensitized with ovalbumin (30 microg i.p.) on days 1 and 6, followed by 5% ovalbumin aerosol challenge on days 11, 12, and 13. A(2A)AR (gene and protein expression), cAMP, and phosphorylated PKA (p-PKA) levels were decreased in A(2A)WT sensitized mice compared with controls. A(2A)KO mice also showed decreased cAMP and p-PKA levels. A(2A)WT sensitized and A(2A)KO control mice had increased gene and protein expression of NADPH oxidase subunits (p47phox and gp91phox) compared with the controls. Tracheal relaxation to specific A(2A)AR agonist, 4-[2-[[6-amino-9-(N-ethyl-beta-d-ribofuranuronamidosyl)-9H-purin-2-yl]amino]ethyl]benzenepropanoic acid hydrochloride (CGS 21680), decreased in A(2A)WT sensitized mice compared with the controls, although it was absent in A(2A)KO mice. Pretreatment with NADPH oxidase inhibitors apocyanin/diphenyliodonium reversed the attenuated relaxation to CGS 21680 in A(2A)WT sensitized tracheas, whereas specific PKA inhibitor (9S,10S,12R)-2,3,9,10,11,12-hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-i] [1,6]benzodiazocine-10-carboxylic acid hexyl ester (KT 5720) blocked CGS 21680-induced relaxation. Tracheal reactive oxygen species (ROS) generation was also increased in A(2A)WT sensitized and A(2A)KO control mice compared with the controls. In conclusion, this study shows that A(2A)AR deficiency causes increased NADPH oxidase activation leading to decreased tracheal relaxation via altered cAMP-PKA signaling and ROS generation.

Interleukin (IL)-1 regulates ozone-enhanced tracheal smooth muscle responsiveness by increasing substance P (SP) production in intrinsic airway neurons of ferret.

Exposure to ozone induces airway hyperresponsiveness (AHR) mediated partly by substance P (SP) released from nerve terminals of intrinsic airway neurons. Our recent studies showed that interleukin (IL)-1, an important multifunctional proinflammatory cytokine, increases synthesis and release of SP from intrinsic airway neurons. The purpose of this study is to investigate the possible involvement of endogenous IL-1 in modulating neural responses associated with ozone-enhanced airway responsiveness. Ferrets were exposed to 2ppm ozone or filtered air for 3h. IL-1 in the bronchoalveolar lavage (BAL) fluid was significantly increased in ozone-exposed animals and responses of tracheal smooth muscle to methacholine (MCh) and electrical field stimulation (EFS) were elevated significantly. Both the SP nerve fiber density in tracheal smooth muscle and the number of SP-containing neurons in airway ganglia were significantly increased following ozone exposure. Pretreatment with IL-1 receptor antagonist (IL-1 Ra) significantly diminished ozone-enhanced airway responses to EFS as well as ozone-increased SP in the airway. To selectively investigate intrinsic airway neurons, segments of ferret trachea were maintained in culture conditions for 24h to eliminate extrinsic contributions from sensory nerves. The segments were then exposed to 2ppm ozone in vitro for 3h. The changes of ozone-induced airway responses to MCh and EFS, and the SP levels in airway neurons paralleled those observed with in vivo ozone exposure. The ozone-enhanced airway responses and neuronal SP levels were inhibited by pretreatment with IL-1 Ra. These findings show that IL-1 is released during ozone exposure enhances airway responsiveness by modulating SP expression in airway neurons.

Nerve growth factor-enhanced airway responsiveness involves substance P in ferret intrinsic airway neurons.

Nerve growth factor (NGF), a member of the neurotrophin family, enhances synthesis of neuropeptides in sensory and sympathetic neurons. The aim of this study was to examine the effect of NGF on airway responsiveness and determine whether these effects are mediated through synthesis and release of substance P (SP) from the intrinsic airway neurons. Ferrets were instilled intratracheally with NGF or saline. Tracheal smooth muscle contractility to methacholine and electrical field stimulation (EFS) was assessed in vitro. Contractions of isolated tracheal smooth muscle to EFS at 10 and 30 Hz were significantly increased in the NGF treatment group (10 Hz: 33.57 +/- 2.44%; 30 Hz: 40.12 +/- 2.78%) compared with the control group (10 Hz: 27.24 +/- 2.14%; 30 Hz: 33.33 +/- 2.31%). However, constrictive response to cholinergic agonist was not significantly altered between the NGF treatment group and the control group. The NGF-induced modulation of airway smooth muscle to EFS was maintained in tracheal segments cultured for 24 h, a procedure that causes a significant anatomic and functional loss of SP-containing sensory fibers while maintaining viability of intrinsic airway neurons. The number of SP-containing neurons in longitudinal trunk and superficial muscular plexus and SP nerve fiber density in tracheal smooth muscle all increased significantly in cultured trachea treated with NGF. Pretreatment with CP-99994, an antagonist of neurokinin 1 receptor, attenuated the NGF-induced increased contraction to EFS in cultured segments but had no effect in saline controls. These results show that the NGF-enhanced airway smooth muscle contractile responses to EFS are mediated by the actions of SP released from intrinsic airway neurons.

Interleukin-1beta-induced airway hyperresponsiveness enhances substance P in intrinsic neurons of ferret airway.

Interleukin (IL)-1beta causes airway inflammation, enhances airway smooth muscle responsiveness, and alters neurotransmitter expression in sensory, sympathetic, and myenteric neurons. This study examines the role of intrinsic airway neurons in airway hyperresponsiveness (AHR) induced by IL-1beta. Ferrets were instilled intratracheally with IL-1beta (0.3 microg/0.3 ml) or saline (0.3 ml) once daily for 5 days. Tracheal smooth muscle contractility in vitro and substance P (SP) expression in tracheal neurons were assessed. Tracheal smooth muscle reactivity to acetylcholine (ACh) and methacholine (MCh) and smooth muscle contractions to electric field stimulation (EFS) both increased after IL-1beta. The IL-1beta-induced AHR was maintained in tracheal segments cultured for 24 h, a procedure that depletes SP from sensory nerves while maintaining viability of intrinsic airway neurons. Pretreatment with CP-99994, an antagonist of neurokinin 1 receptor, attenuated the IL-1beta-induced hyperreactivity to ACh and MCh and to EFS in cultured tracheal segments. SP-containing neurons in longitudinal trunk, SP innervation of superficial muscular plexus neurons, and SP nerve fiber density in tracheal smooth muscle all increased after treatment with IL-1beta. These results show that IL-1beta-enhanced cholinergic airway smooth muscle contractile responses are mediated by the actions of SP released from intrinsic airway neurons.

Pulmonary alterations associated with inhalation of occupational and environmental irritants.

Many gases, vapors, or particles found in occupational and/or environmental settings can act as irritants. In the present study, sensory irritants are characterized by the stimulation of neuropeptide release from sensory nerves in the nasal mucosa, while pulmonary irritants are characterized by recruitment of PMN into bronchoalveolar airspaces, elevation of breathing frequency, and neuropeptide release from sensory fibers innervating the epithelium of the conducting airways. A review of data from our laboratory as well as results from others indicate that asphalt fume is a sensory irritant; toluene diisocyanate (TDI), methyl isocyanate, and machining fluid act as both sensory and pulmonary irritants; while cotton dust, agricultural dusts, microbial products, leather conditioner, and ozone exhibit responses characteristic of pulmonary irritants.

Role of intrinsic airway neurons in ozone-induced airway hyperresponsiveness in ferret trachea.

Exposure to ozone (O(3)) enhances airway responsiveness, which is mediated partly by the release of substance P (SP) from airway neurons. In this study, the role of intrinsic airway neurons in O(3)-induced airway responses was examined. Ferrets were exposed to 2 ppm O(3) or air for 1 h. Reactivity of isolated tracheal smooth muscle to cholinergic agonists was significantly increased after O(3) exposure, as were contractions to electrical field stimulation at 10 Hz. Pretreatment with CP-99994, a neurokinin type 1 receptor antagonist, partially abolished the O(3)-induced reactivity to cholinergic agonists and electrical field stimulation. The O(3)-enhanced airway responses were present in tracheal segments cultured for 24 h, a procedure shown to deplete sensory nerves while maintaining viability of intrinsic airway neurons, and all the enhanced smooth muscle responses were also diminished by CP-99994. Immunocytochemistry showed that the percentage of SP-containing neurons in longitudinal trunk and the percentage of neurons innervated by SP-positive nerve fibers in superficial muscular plexus were significantly increased at 1 h after exposure to O(3). These results suggest that enhanced SP levels in airway ganglia contribute to O(3)-induced airway hyperresponsiveness.

Projections and pathways of VIP- and nNOS-containing airway neurons in ferret trachea.

Airway ganglia in ferret trachea are organized into two identifiable plexuses, the longitudinal trunk (LT) ganglia and the superficial muscular plexus (SMP) ganglia. LT ganglia contain cholinergic neurons and neurons in SMP ganglia synthesize vasoactive intestinal peptide (VIP) and nitric oxide (NO). Both plexuses of ganglia project to airway smooth muscle, but communications between LT and SMP ganglia have not been extensively studied. Ganglia of the LT were injected with neural tracers, either rhodamine- or biotin-labeled dextran amines, and colocalized with VIP or neuronal NO synthase (nNOS) by immunocytochemistry. Anterograde tracers were found in nerve fibers that formed basket-like complexes associated with neurons of LT and SMP ganglia and were observed in the nerve fibers of smooth muscle and tracheal plexus. Some VIP or nNOS positive neurons in the SMP ganglia contained retrogradely transported tracer. The findings suggest that neurons in the SMP and LT ganglia communicate in a reciprocal fashion and that VIP and nNOS neurons both receive and send neural inputs to neurons of LT, presumably cholinergic, ganglia. These connections probably contribute to the integration of neural control of airway and vascular smooth muscle tone and glandular secretion in the airways.

Effect of ozone treatment on airway reactivity and epithelium-derived relaxing factor in guinea pigs.

Ozone (O(3)) is toxic to respiratory epithelium and causes airway inflammation and hyperreactivity. To evaluate the role of the epithelium in the development of hyperreactivity, we examined in guinea pigs the effects of inhaled O(3) (3 ppm for 1 h; 0-24 h after exposure) on 1) reactivity to inhaled methacholine (MCh), 2) reactivity of the isolated, perfused trachea (IPT) to MCh, 3) epithelium-derived relaxing factor (EpDRF)-mediated relaxations of IPT induced by mucosal hyperosmolar solutions, 4) neurogenic contraction and relaxation responses, 5) transepithelial potential difference, and 6) microscopic analysis of nitrotyrosine immunofluorescence, substance P fiber density, and tracheal morphology. At 0 h, O(3) caused hyperreactivity to inhaled MCh and mucosally but not serosally applied MCh in IPT (only in the presence of the epithelium) and a decrease in transepithelial potential difference. Inhibition of EpDRF-induced relaxation responses occurred at 2 h. All of these changes returned to control by 12 to 18 h. O(3) had no effect on neurogenic responses. Nitrotyrosine immunofluorescence appeared in the trachea at 0 h in detached epithelial cell ghosts and in intrapulmonary airways by 6 h. Substance P fiber density was elevated in smooth muscle at 0 and 18 h but not in epithelium or lamina propria of intrapulmonary and extrapulmonary bronchi. Loss of cilia and mucosubstances in the mucosa occurred at 0 h; the epithelium became markedly attenuated over 12 to 24 h. A reversible increase in epithelial permeability and a decrease in EpDRF production may contribute to O(3)-induced hyperreactivity to MCh.

Toluene diisocyanate enhances substance P in sensory neurons innervating the nasal mucosa.

Inhalation of irritants, such as toluene diisocyanate (TDI), stimulates substance P (SP) release from peripheral processes of sensory neurons innervating the airways. The purpose of this study was to determine if TDI inhalation affects intraneuronal levels of SP and preprotachykinin (PPT) messenger RNA (mRNA) in the sensory neurons of the trigeminal ganglion (TG) which innervate the nasal epithelium. The nasal cavity of Fisher-344 rats was instilled with rhodamine-labeled latex microspheres. Ten days later, the rats were exposed to 60 ppb of 2,4-2,6-TDI vapor for 2 h. The TG were removed 1, 12, 24, 48, 72, and 96 h after TDI treatment and prepared for SP immunocytochemistry and PPT in situ hybridization. SP nerve fiber density in nasal epithelium was significantly increased 12, 24, and 48 h after TDI exposure. The proportion of microsphere-labeled cell bodies expressing high levels of SP immunoreactivity was decreased at 24 h but was increased above controls at 48 and 72 h. The proportion of microsphere-labeled cell bodies expressing high levels of PPT mRNA was increased above control levels at 24 and 48 h. The percentage of leukocytes observed in nasal lavage fluid was significantly increased 12, 24, 48, and 72 h after inhalation. These studies indicate that SP production in TG neurons projecting to the nasal epithelium is transiently increased after TDI exposure, suggesting that TDI inhalation not only causes SP release but also increased intraneuronal neuropeptide levels. Increased neuronal SP levels may be involved in maintaining neurogenic inflammation or the development of airway hyperresponsiveness.

Innervation of tracheal epithelium and smooth muscle by neurons in airway ganglia.

The neurochemical profiles of neurons in ferret tracheal ganglia has been characterized, but their projections to smooth muscle and epithelium in ferret trachea has not been examined. The purpose of this study is to determine the location of cell bodies that project VIP-, SP-, and NPY-containing fibers to the ferret tracheal smooth muscle and epithelium. Segments of ferret trachea were cultured for 0, 1, 3, or 7 days, some in the presence of 3 microm capsaicin. VIP, SP, or NPY nerve fiber density was measured using morphometric procedures. A retrograde tracer, rhodamine-labeled microspheres, identified neurons projecting to the epithelium. The density of SP fibers in the epithelium was reduced after culture, but VIP innervation was not different. In tracheal smooth muscle, the density of VIP- and SP-IR fibers was not different during the culture period, but NPY fiber density was reduced at all culture times. Capsaicin treatment did not affect nerve fiber density in the tracheal smooth muscle but produced a significant reduction in the density of epithelial VIP- and SP-IR nerve fibers after 1 day. Rhodamine-labeled microspheres were identified in VIP-containing nerve cell bodies of the ferret tracheal plexus. VIP innervation to the airway epithelium in ferret originates both from cell bodies in airway ganglia and cell bodies in sensory ganglia. The pathway from airway ganglia suggest the existence of a local reflex mechanisms initiated by epithelial irritation.

Characterization of neural control and contractile function in airway smooth muscle of the ferret.

Several recent studies have characterized neuroanatomical and neurophysiological aspects of ferret airways, but regional differences in reactivity and cholinergic, adrenergic and non-adrenergic-non-cholinergic (NANC) neural responses have not been examined. The aim of this study was to characterize the contractile and relaxant response elicited by electrical field stimulation (EFS), and the contractions induced by cholinergic agonists in isolated ferret tracheal and bronchial preparations. EFS produced frequency-dependent contractions and relaxations. Contractions in both the trachea and bronchi were inhibited by atropine and potentiated by neostigmine. Tracheal relaxations were found to be entirely adrenergic, but bronchial relaxations were mediated by a combination of adrenergic and inhibitory NANC (i-NANC) innervations. Trachea and bronchi were more sensitive to methacholine (MCh) and carbachol than to acetylcholine (ACh); middle tracheal segments being more sensitive to ACh than distal segments, however, in the presence of neostigmine ACh potency was equal in both segments. The results suggest that regional differences exist in cholinergic responsiveness of ferret trachea and bronchi, resulting from differences in ACh degradation.

Enhancement of nitric oxide production by pulmonary cells following silica exposure.

In vivo exposure of rat lungs to crystalline silica either by intratracheal instillation or by inhalation results in an increase in mRNA levels for inducible nitric oxide synthase (iNOS) in bronchoalveolar lavage cells (BALC), elevated nitric oxide (.NO) production by BALC, and an increase in .NO-dependent chemiluminescence (CL) from alveolar macrophages (AM). Induction of iNOS message occurs in both AM and polymorphonuclear leukocytes (PMN) harvested from silica-exposed lungs but is not significantly elevated in lavaged lung tissue. In vitro exposure of AM to silica does not stimulate .NO production or enhance iNOS message. However, treatment of naive AM with conditioned media from BALC harvested from silica-exposed rats does increase iNOS message and .NO production by these AM. The potency of this conditioned medium is dependent on interaction between AM and PMN. In the rat model, a relationship exists between the ability of various dusts to cause PMN recruitment or protein leakage into the alveolar space and the induction of iNOS message in BALC, i.e., silica > coal mine dust > carbonyl iron > titanium dioxide. Similarly, a comparison of BALC from a healthy volunteer, a silica-exposed coal miner with a normal chest radiograph, and a silica-exposed coal miner with an abnormal chest radiograph shows a correlation between pathology and both the level of iNOS message in BALC and the magnitude of .NO-dependent CL from AM. These data suggest that .NO may play a role in silicosis and that human pulmonary phagocytes exhibit enhanced .NO production in response to an inflammatory insult.

Effects of silica exposure on substance P immunoreactivity and preprotachykinin mRNA expression in trigeminal sensory neurons in Fischer 344 rats.

Trigeminal sensory neurons innervate the nasal cavity and may release substance P (SP) upon exposure to inhaled irritants. The purpose of this study was to determine if silica dust, an occupational irritant causing inflammation, activates sensory neurons supplying the nasal cavity. Male Fischer 344 rats were placed in inhalation chambers and exposed daily to 2 mg/m3 of fresh silica (average diameter 1 microm) for 6 mo. Following exposure, the trigeminal ganglia (TG) were removed and prepared for SP immunocytochemistry and for preprotachykinin (PPT) autoradiographic in situ hybridization. The SP-like immunofluorescence in TG neurons was subjectively categorized as high, moderate, or low (background) intensity. In situ hybridization autoradiographs were quantified on the basis of grain density using digital imaging analysis. The SP immunoreactivity and PPT mRNA expression in the TG neurons were significantly increased after silica inhalation. The proportion of highly positive SP-immunoreactive neurons shifted from 1.30 +/- 0.58% in controls to 11.30 +/- 1.15% after silica treatment. The neurons exhibiting high grain density for PPT mRNA increased from 1.50 +/- 0.87% in controls to 11.67 +/- 0.58% in the silica group. Thus, inhalation of silica causes upper airway irritation resulting in increased levels of immunoreactive neuronal SP and PPT mRNA. These findings suggest that silica activates sensory pathways that may be involved in nasal inflammation.

N-methyl-D-aspartate receptors are expressed by intrinsic neurons of rat larynx and esophagus.

Overactivation of N-methyl-D-aspartate receptors (NMDAR), a mechanism of central neurotoxicity, has recently been shown to increase airway responsiveness in rat lungs. NMDAR have not been localized in the airways, but neurons of the myenteric plexus in the rat express mRNA for NMDAR. Furthermore, a population of glutamate-containing cell bodies in the nucleus ambiguus projects to the rat larynx. On this basis, we hypothesized that some postganglionic parasympathetic neurons of the larynx, trachea and esophagus may express NMDAR. Sections of rat larynx, trachea and esophagus were immunocytochemically labeled for NMDAR subtype 2B using a specific antibody. NMDAR immunoreactivity was observed in cell bodies of individual neurons located in the submucosa and on the external surface of skeletal muscle in the larynx and also in neurons of the esophageal plexus. All NMDAR-positive nerve cell bodies also contained immunoreactivity for vasoactive intestinal peptide (VIP) and some were immunoreactive for nitric oxide synthase (NOS). None of the cell bodies of the tracheal plexus contained NMDAR immunoreactivity. The findings demonstrate that NMDAR are expressed in neurons of the rat larynx and esophagus and their activation may be associate with VIP or NO release.

Identification and neuropeptide content of trigeminal neurons innervating the rat nasal epithelium.

Trigeminal ganglia provides sensory innervation to the rat nasal cavity. The purpose of this study was to identify the location and characterize the neuropeptide content of trigeminal neurons that project specifically to the rat nasal epithelium. The right nasal cavity was instilled with 4 microliters of rhodamine-labelled latex microspheres. Seven, 10 or 14 days after tracer instillation, both trigeminal ganglia were removed and prepared for substance P and calcitonin gene-related peptide immunocyto-chemistry. neurons labelled with microspheres were located in the trigeminal ganglia at the division of the ophthalmic and maxillary nerves and were only found on the side ipsilateral to the instillation. The percentage of labelled cell body profiles in the right trigeminal ganglia averaged 1.61 +/- .04% at seven days. 1.54 +/- .01% at 10 days, and 1.65 +/- .02% at 14 days after instillation. These values were not statistically different, but the fluorescence intensity in labelled neurons was increased after 14 days. The right trigeminal ganglia contained a mean of 166 +/- 13.81 labelled cell body profiles representing 1.60 +/- .09% of the total. Of the labelled profiles, 81.6 +/- 3.27% were immunoreactive for substance P and 35.2 +/- 4.00% for calcitonin gene-related peptide. Sections of nasal mucosa showed that the microspheres were localized only in the epithelial layer and did not enter the lamina propria indicating that the microsphere transport was confined to nerve fibres present in the epithelial layer. These studies have identified specific neurons in the trigeminal ganglia that project to the nasal epithelium. Most of the neurons retrogradely labelled from the nasal cavity contained substance P but some contained calcitonin gene-related peptide, suggesting that the nasal epithelium is predominantly targeted by substance P-containing neurons.

Glutamate toxicity in the lung and neuronal cells: prevention or attenuation by VIP and PACAP.

VIP, which has been demonstrated to reduce or prevent oxidant injury in the lungs and other organs, is shown here to protect against excitotoxic injury of the lung and excitotoxic death of cortical neuronal cells in primary culture. Glutamate killing of neuron-like PC-12 cells, attributable to oxidant stress rather that to excitotoxicity, is also reduced or prevented by VIP and by the closely related peptide PACAP. The exact mechanisms of this protection remain to be determined, but appear to include antioxidant and anti-apoptotic actions, and suppression of glutamate-induced upregulation of its own receptor. Both VIP and PACAP offer the promise of novel and nontoxic means of defending against NMDA and glutamate toxicity.

Rat alveolar macrophages express preprotachykinin gene-I mRNA-encoding tachykinins.

Although the tachykinins substance P (SP) and neurokinin A have been largely localized to neurons, eosinophils have also been shown to express these peptides. Our aim was to determine whether rat alveolar macrophages (AM) express preprotachykinin gene-I (PPT-I) mRNA that encodes these tachykinins and to examine expression during inflammation. PPT-I mRNA was detected by reverse transcription (RT)-polymerase chain reaction (PCR) in AM and brain (control) but not in peritoneal macrophages. Northern analysis showed that PPT-I mRNA was induced two- to fourfold by in vivo treatment of rats with intratracheal lipopolysaccharide (LPS) and in vitro after 4 h of exposure to LPS. This increase was inhibited by dexamethasone. In situ RT-PCR and immunocytochemistry further confirmed that AM express PPT-I mRNA and SP-like immunoreactivity, respectively, which was enhanced by LPS treatment. A 1.3-kb transcript consistent with PPT-I mRNA was detected by Northern analysis of bronchoalveolar lavage neutrophils. Therefore, rat AM express PPT-I mRNA that is upregulated in AM by LPS and is attenuated by dexamethasone. PPT-I mRNA was also detected in lung neutrophils.