Intranasal Histamine, Methacholine, and Bradykinin Challenge in Children With and Without Allergy

Andrew G. Vayonis, M.D.,* James T. Seroky, M.A.,t William J. Doyle, Ph.D.,t David P.Skoner, M.D.,* and Philip Fireman, M.D.*

ABSTRACT

Property ofymptoms of allergic rhinitis include rhinorrhea, sneez-

ing, nasal congestion, and nasal pruritis. These sympIn this study, allergic and non allergic children (7-13 years) toms, along with objective findings of increased nasal were challenged with increasing doses (0.0], 0.], 1.0 mg) of resistance, eustachian tube dysfunction, and increased histamine, methacholine, and bradykinin at different study sesnasal secretions, are commonly observed following sions. The effects of these challenges on nasal patency, seprovocative nasal challenges using specific allergens in cretion weight, sneezing, nasal symptoms, eustachian tube sensitized subjects.I-3 Also, assays of nasal lavage fluids function, middle-ear pressure, and pulmonary function were after allergen challenge have documented elevations of documented. Although intranasal challenges with the three several inflammatory mediators including histamine, substances provoked nasal symptoms and secretion, only histabradykinin, the leukotrienes, and the prostaglandins.4-8 mine challenge provoked sneezing and resulted in significant Other studies have evaluated the isolated effects of inchanges in the measures of nasal patency and eustachian tube tranasal provocative challenge with these inflammatory function. None of the substances provoked changes in middle mediators on nasal symptoms, signs, and pathophysioloear pressures or pulmonary function. Allergic status had a sig. . gles. 9-23 The resu 1 0f a comparatIve study of adult subts nificant effect on secretion weight and symptoms of congestion jects with and without allergic rhinitis documented mediand rhinorrhea, most consistently documented as a greater reator specificity with respect to the signs and symptoms sponse of the allergic subjects to histamine challenge. In genOceanSide Publicationsby intranasal challenges using histamine, provoked eral, the results of this study in children are similar to those methacholine, bradykinin, and prostaglandins D2 and previously described for challenges with these substances in IP: 152.118.148.218 F2alpha.23 adults. (American Journal of Rhinology 9:1-7,1995) With the exception of a report by Tomanaga and colleagues,21 all of the provocative mediator challenges were performed on adult subjects. The present study was conducted to extend our understanding of the effects of topical intranasal challenge with inflammatory substances to children with and without allergic rhinitis. The primary hypothesis is that the mediator specificities in provoked signs and symptoms documented in adults also characterize the responses of children. A secondary hypothesis is that, like allergic adults, children with documented allergic rhinitis exhibit a nasal hyperresponsiveness to these challenges when compared with non-allergic children. To allow for comparisons with the adult data reported previously, the challenge substances used were histamine, methacholine, and bradykinin.23 Both subjective and objective response measures including symptom scores,

From the Department of Pediatrics, Division of Allergy, Immunology, and Rheumatology* and the Department of Otolaryngology, t Children's Hospital of Pittsburgh and the University of Pittsburgh School of Medicine Supported in part by a grant from the National Institutes of Health No. AI/9262 and General Clinical Research Centers Grant MO/RR 00084 Address correspondence and reprint requests to Philip Fireman, M.D., Department of Pediatrics, Division of Allergy, Immunology, and Rheumatology, Children's Hospital of Pittsburgh, 3705 Fifth Avenue at DeSoto Street, Pittsburgh, PA ]5213

American Journal of Rhinology

Nutley, NJ), or bradykinin (Peninsula Lab, Belmont, CA). On the day of challenge, these were prepared as 1 mg/mL stock solutions in diluent (Dulbecco phosphatebuffered saline-bradykinin; normal saline-histamine and methacholine). The solution for histamine was calculated from the weight of the salt. The pH was adjusted to 7.4 METHODS with phosphate buffer for bradykinin and to 7.0 with Study Population NaOH for histamine and methacholine. Ten-fold diluubjects between the ages of 7 and 13 and in good tions of the stock solution with diluent were made to health were recruited from the patients of the Asthma yield the challenge doses. All 32 subjects completed the and Allergic Disease Center of Children's Hospital of histamine and methacholine challenge sessions and 25 Pittsburgh and from residents of the surrounding commu(12 allergic rhinitis, 13 nonallergic) subjects completed nity. Parents of potential subjects were interviewed and the bradykinin challenge session. provided a detailed history for their child regarding alFor each session, subjects and their parent/guardian relergy, asthma, nasal disease, otologic disease, symptoported to the laboratory approximately 30 minutes before matic upper respiratory tract infections within the past 4 challenge to allow for acclimation. After examination of ears, nose, throat, and lungs, baseline measurements of weeks, and any other illnesses. A general physical examination was performed on the child. Subjects were exnasal patency by anterior rhinomanometry, eustachian cluded from participation if presenting with signs and tube function by sonotubometry, middle-ear pressure by symptoms of allergic rhinitis, upper respiratory tract intympanometry, and pulmonary function by spirometry fection, middle ear disease, or if they had a positive hiswere made, and prechallenge symptoms were scored. PropertyThen, all subjects were challenged intranasally at 15of tory of any chronic medical condition, otologic disease, asthma, or upper respiratory tract infection within the minute intervals, with the diluent followed by increasing past 4 weeks. The parent or guardian of enrolled subjects doses of the challenge substance (0.01, 0.1, 1.0 mg). provided a written informed consent, and the child proDoses were chosen based on our previous experience with these substances in adults.23 For delivery, 0.5 mL of vided a verbal assent. All enrolled subjects were evaluated for allergic rhinithe prepared solution was placed in a nebulizer (DeViItis. A positive diagnosis of allergic rhinitis included a hisbiss atomizer model 251) and administered into one side tory of at least 2 years of symptoms consistent with nasal of the nasal cavity under pressure (Pulmo-Aide pump, DeVilbiss Co.) during deep inhalation while the contralatallergy and a positive puncture and/or intradermal skin test to at least one of the common aeroallergens including eral nostril was manually occluded. The nebulizer pump ragweed mixture, tree mixture, grass mixture, and dust was activated at the onset of a deep inspiratory maneuver mites. Subjects assigned to the nonallergic group did not and deactivated at the termination of the maneuver (aphave a positive history of allergy or positive skin tests to proximately 3 to 5 seconds). The procedure was repeated the panel of allergens tested. Seventeen children with if the total volume of the challenge substance was not delivered in the first attempt. After administration of each allergic rhinitis (8 males; aged 7 to 13 years, average 10.8::1.7 years) and 15 nonallergic controlOceanSide Publications (8 males, challenge dose, the tests of nasal patency, eustachian tube function, middle-ear pressure, and pulmonary function aged 8 to 13 years, average 10.5::1.5 years) subjects were studied. were repeated; symptoms were scored, and the number of IP: 152.118.148.218 sneezes and coughs were counted by the technician. ProProtocol voked secretions were actively expelled into tissues and he enrolled subjects were scheduled for three chalsealed in plastic bags of known weight. lenge sessions with a minimum between-session Specific Methods interval of 5 days. The order sequence of the challenge sessions was histamine, methacholine, and bradykinin. asal airway patency was evaluated for the right, left, Subjects, but not laboratory personnel, were blinded to and total nasal cavity by anterior rhinomanometry, the nature of the challenge substance and its expected efusing a microcomputer assisted rhinomanometric system fects. The allergic subjects were studied "out of season" developed in our laboratory and described previously.23 when they were asymptomatic. No subject was permitted Data were collected during a 20-second period while the to have used antihistamines, decongestants, or nasal crosubject breathed in a relaxed manner. The variable chosen molyn for 72 hours, or topical nasal steroids for 2 weeks. for presentation was the total nasal conductance recorded Astemizole, loratadine, and terfenadine had not been during inspiration with units of liters per second per cenused by any of the subjects in the study. timeter of water. At each session one of three challenge substances was Eustachian tube function was assessed by sonotubomeadministered: histamine dihydrochloride (lCN, Irvine, try using a computerized instrument developed in our California), methacholine (Provocholine) (Roche Lab, laboratory and described previously.23Each test consisted nasal secretion weights, number of sneezes and coughs, eustachian tube function, middle ear pressure, pulmonary function, and nasal conductance were included in the challenge protocol.

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these summary data by a two-way analysis of variance of four attempts to dilate the tube by swallowing. Test repartitioned by challenge substance and allergic status, and sults were classified as yes/no categories based evaluated at P < 0.05. If a significant effect was observed, on whether or not the tube opened on any of the four the Least Significant Difference method was used to deterattempts. mine the nature of the difference. In the presentation, the Pulmonary function was evaluated with spirometry and convention meanstandard deviation is used throughout. a commercial instrument (Multi Spiro-PC). Results were expressed as the per cent predicted forced expiratory volRESULTS ume in 1 second (%FEV1) referenced to a panel of normal subjects. Response Dose Functions Middle ear pressure was measured by tympanometry asal conductance provides a quantitative measure of using a standardized clinical tympanometer (Model No. nasal patency. Figure 1 shows the average values of TA- 7A, automatic impedance meter, Teledyne Avionics, conductance as a function of dose for each of the chalCharlottesville, VA). lenge substances. There were no significant differences Nasal secretion weight was determined by providing between sessions in the average value of nasal conducthe subjects with preweighed tissues and instructing them tance recorded on baseline testing. Neither methacholine to actively expel their nasal secretions into the tissues or bradykinin elicited a dose-dependent decrease in conbefore performing the rhinomanometric testings and at ductance. However, the results for the challenge with hisother times as required. Expended tissues were then tamine showed a progressive decrease in conductance sealed in a plastic bag of known weight. At the end of the with increasing dose. There, the difference between the session, the bag was weighed, and secretion weight was average nasal conductance recorded following the diluent calculated by subtraction of the weights of the tissues and Property challenge and that recorded following the 1.0 mg chalof plastic bag. lenge was statistically significant. Symptoms were scored by the subjects on a 0 to 3 scale Figure 2 shows the average values of the sneeze count (0 = no symptoms, 1 = mild symptoms, 2 = moderate as a function of dose for the three challenge sessions. symptoms, 3 = severe symptoms). Specific symptoms Challenges with bradykinin and methacholine did not listed on the data sheet were nasal congestion, rhinorrhea, provoke increasing sneezing. However, histamine chalheadache, sore throat, and watery/itchy eyes. These lenge resulted in a dose-dependent increase in sneezing, symptoms were chosen to correspond to those reported with a significant difference between the average sneeze previously for mediator challenges in adults. The number count recorded after the 1 mg challenge dose and that of sneezes and coughs was also recorded by the technirecorded after diluent challenge. cian after administration of each challenge dose. Subjects Challenges with all three substances showed no signifwere permitted to comment on any other symptoms not icant dose-dependent increase in the rhinorrhea symptom specifically listed. These other symptoms were recorded score. on the data sheets but were not scored. Histamine, but not bradykinin or methacholine, challenge resulted in a dose-dependent increase in nasal conStatistical Methods gestion symptoms. n the analysis, secretion weight, conductance, sneeze

count, and nasal symptoms were considered to be con0.50 tinuous variables. The results for eustachian tube function IP: 152.118.148.218 and middle ear pressure were treated as dichotomous 0.45 ~ 0.40 variables. To avoid complications associated with multiple comparisons, a positive response to challenge with 0.35 any of the substances was defined as a statistically signif0.30 d icant difference between the value of the response vari0.25 able following diluent challenge and that following the 1 ~ 0.20 mg challenge dose. For continuous variables, this com0.15 parison was made using a paired "t" test evaluated at 0.10 P < 0.05. For the dichotomous variables, frequency data ~ 0.05 were compared using the McNemar test for significance 0.00 fIASE 0 0.01 0.1 of changes. To determine the significance of differences in the reo-tALLENGE DOSE (mg) sponses between allergic and non-allergic subjects and Figure 1. Average nasal conductance as a function of dose for among challenge substances, a summary response measure was defined for each continuous variable. This conhistamine (plus), methacholine (diamond), and bradykinin (trisisted of summing the values of the measured variable angle). Vertical bars indicate the 95% confidence interval about the average values of the histamine data. across active doses (0.01, 0.1, and 1.0 mg) and analyzing

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10 9

8 7 6
5

Analysis of Variance two-way analysis of variance was performed on the summary response variables to evaluate the signifi:3 ~ cance of the contributions of challenge substance and al2 lergic status to the measured responses. Table II presents the F values and level of significance by source for the 0 BASE 0 0.01 0.1 summary response measures. A significant contribution of challenge session to total variance was documented for sneeze count, secretion weight, and symptoms of nasal Figure 2. Number of sneezes as a function of dose for histacongestion. This is interpretable as differences in the mine (plus), methacholine (diamond), and bradykinin (trianmagnitudes of those responses provoked by the three gle). Vertical bars indicate the 95% confidence interval about challenge substances. A significant contribution of allerthe average values of the histamine data. gic status to total variance was documented for secretion weight and symptoms of nasal congestion and rhinorrhea. This implies a difference in the response to challenge Cough was not provoked by any of the challenge substances in either the allergic or nonallergic subjects. Propertywith these substances in allergic and non-allergic subjects. of Table III reports the average values and standard deviaSymptoms of sore throat, headache, watery eyes, and tions of these response variables following challenge itchy eyes were reported by a number of subjects followwith each of the three substances in the allergic and noning challenges with histamine and methacholine, but not allergic subgroups. Those data show that for the three bradykinin. The "other" symptom that was reported most variables with a significant effect of challenge session frequently following challenge with all three substances documented by the analysis of variance, the primary difwas mild burning in the nose. (Table I). For unclear reaference is a greater response to histamine challenge when sons, this appeared to be more common in the non-allercompared with the lesser and approximately equal regic group using histamine and methacholine as compared sponses to methacholine and bradykinin challenge. This to a greater frequency in the allergic group using bradydifferential response with respect to challenge substance kinin. Watery eyes appeared to be more common in the alcharacterized both the allergic and non allergic sublergic group with histamine and methacholine challenges. groups. For the three variables with a documented effect On baseline testing, eustachian tube function was of allergic status, the allergic group had a greater rejudged to be abnormal in approximately 30% of the ears sponse when compared with the nonaJlergic group folfor the histamine and methacholine challenge sessions lowing challenges with all three substances. and in approximately 50% of the ears for the bradykinin OceanSide challenge session. A dose-dependent increase in the fre- Publications DISCUSSION quency of abnormal function tests was observed for the IP: 152.118.148.218 allergen challenge in adults has been used to ntranasal histamine (maximum increase = 28%) and bradykinin diagnose nasal allergies, evaluate the efficacy of ther(maximum increase = 15%), but not the methacholine apies, and provide insights into the pathogenesis of the challenge sessions. However, the difference between the frequencies recorded following the diluent challenge and the 1.0 mg challenge were only significant for the histaTABLE I mine challenge session. There was no difference between the response of allergic and non-allergic subjects to histaNumber of Subjects in the Allergic (AR) and mine challenge. Nonallergic (NAR) Subgroups Reporting Average values of middle-ear pressure were plotted as Other Symptoms During the Course of the a function of challenge dose for each of the three mediaThree Challenge Sessions tors. None of the challenges resulted in dose-dependent Histamine Methacholine Bradykinin changes in the average middle-ear pressure. AR AR NAR AR NAR NAR Average baseline %FEV1 ranged from 85% to 128% Sore throat 1 1 0 2 o 0 with no statistically significant difference between the alHeadache 4 2 1 1 o 0 Watery eyes 4 1 6 1 o 0 lergics and controls. No dose-dependent decrease in the Itchy eyes 2 2 2 o 0 0 average value of this variable was observed for any of the Nasal burning 2 2 6 5 4 0 challenge sessions. The data for each subject were exam4

ined for changes from baseline. A decrease from baseline FEV1 of greater than 20% was observed in one allergic subject following histamine challenge, and in a second following methacholine challenge. No wheezing was auscultated, and neither subject was symptomatic.

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TABLE II F Values and Level of Significance 0 For the Analysis of Variance of the Continuous Variables by Source Substance df Conductance Secretion weight Sneeze count Congestion score Rhinorrhea score
df 2.4

Allergic Status 1.6 6.4 0.3 10.7 6.3 (0.21) (0.01) (0.56) 0.01) (0.01) 1.0 3.3 0.2 0.8 0.8

Interaction (0.38) (0.04) (0.84) (0.93) (0.93)

213
10.5 23.3 4.9 1.6 (0.10)* 0.01) 0.01) 0.01) (0.20)

* = Probability
challenges with these substances do not provoke late redisease expression.2-7 Similarly, challenges with substances elaborated during nasal allergic reactions have actions and are primarily associated with the development of acute, self-limiting nasal symptoms and pathobeen used to define the symptomatic and pathophysiologic effects of the substances in isolation, predict the efphysiologies.23 Challenge doses were chosen so as to duplicate the lower range of doses used in previous adult fects of treatment with substance specific interventions, and diagnose altered states of nasal responsiveness.9-23 studies in our laboratory. Allergic and nonallergic subPropertyjects were included to test the hypothesis of a nasal of Regarding the latter, numerous studies have been conducted using a variety of substances with suspected hyper-responsiveness to challenge in the allergic subgroup. Where possible, objective measures of the nasal inflammatory potential. The results have documented response were used to supplement the subjective reportsubstance specificity in eliciting symptomatic and pathophysiologic responses as well as an enhanced responsiveing of symptoms by the child. The results of this study show that intranasal challenge ness of allergic rhinitis patients to intranasal challenges.23 of children using the described methods, substances, and The majority of these studies were conducted on adult doses is relatively safe. No child experienced any adverse subjects. However, allergic rhinitis can occur at any age, or unexpected reactions to any of the challenges. Generincluding early infancy, and most patients develop their alized, nonspecific symptoms were not volunteered by symptoms during childhood.24,25It is not known if the efthe subjects, and the reported symptoms were confined fects of challenges with these substances documented for mostly to the nose and throat. No challenge session was adults can be extrapolated to children. discontinued because of distress experienced by the The present study was designed to address this issue. A child. Possible pulmonary complications of the chalprimary concern in developing the study design was the lenges were monitored by spirometry, and the results safety of performing intranasal challenges in children. showed no consistent changes associated with any of the Consequently, the panel of substances evaluated was limOceanSide Publicationssubstances. A decrease in the FEV1 of 20% ited to bradykinin, histamine, and methacholine because challenge extensive experience in adults showed that intranasal with respect to baseline was observed during 2 of the 79

= Degrees

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TABLEllI

Means and Standard Deviations of the Summary Response Variables for Challenges with Histamine, Bradykinin, and Methacholine in the Allergic (AR) and Nonallergic (NAR) Subgroups Variable Conductance Secretion weight Sneeze count Congestion score Rhinorrhea score Histamine Bradykinin 0.81 1.09 2.1 2.1 0.8 0.5 2.8 1.3 1.5 0.5 0.36 0.71 1.0 1.2 1.1 0.8 2.7 1.9 1.9 1.1 Methacholine 0.94 0.85 2.4 1.8 1.2 1.1 2.0 0.7 1.3 0.5 0.86 0.50 1.2 0.9 1.7 1.4 1.6 1.5 1.6 0.9

AR
NAR
AR

NAR

AR
NAR
AR

NAR
AR

NAR

0.51 0.78 4.9 2.7 11.2 9.3 3.9 2.2 1.9 1.2

0.23 0.44 2.9 1.7 11.2 7.8 2.7 1.6 1.8 1.5

American Journal of Rhinology

sessions. However, neither child was Otitis media with effusion is common in the pediatric symptomatic or had wheezing on auscultation of the population and a variety of evidences support a role for lungs, and the abnormality rapidly reversed without interallergy in the pathogenesis of that disease. For example, vention. Possible otologic complications of the chalTomonaga and colleagues reported a significantly higher lenges were monitored by tympanometry that did not frequency of allergic rhinitis in children dia~nosed with document any effects on middle ear pressures?3 otitis media as compared to control children. 1 They sugIn general, the responses observed in this population gested that the relationship is mediated by the eustachian were similar to those previously reported for adults chaltube dysfunction that accompanies a nasal allergic reaclenged with these three substances. Specifically, intion. Indeed, a number of investigators reported the detranasal histamine challenges provoked the greatest revelopment of tubal dysfunction following natural and exsponse magnitudes. The nasal responses to histamine perimental allergen challenges in sensitized subjects, and included significant increases in sneezing and symptoms others provoked tubal dysfunction in adults following challenge with specific inflammatory substances. 1,20,23 In of nasal congestion and decreases in the objective measure of nasal patency. The magnitude of these effects was one study of children, intranasal histamine challenge dose related. The symptoms of rhinorrhea provoked by by the disc method resulted in eustachian tube dysfunchistamine challenge showed a non-significant dosetion?O The results of the present study confirm that obserrelated increase, but the objective measure of secretion vation. Specifically, histamine challenge resulted in a production was significantly greater than that recorded dose-dependent increase in the frequencies of eustachian following challenges with the other two substances. tube dysfunction. Bradykinin showed a similar doseBradykinin challenges resulted in dose-dependent inrelated effect, although not of sufficient magnitude to creases in symptoms of rhinorrhea and congestion, alachieve statistical significance. Methacholine challenge Propertydid not affect tubal function. However, as described of though these changes were not statistically significant. Bradykinin challenge did not provoke sneezing and had above and reported for adult challenges, these changes in no significant effect on the objective measure of nasal patubal function were not accompanied by effects on midtency. Secretion weights were significantly less than dle ear pressure. those recorded during histamine challenge and equal to In conclusion, the responses to challenges with histamine, bradykinin, and methacholine are similar for chilthose observed following methacholine challenge. Of the measured functions, only symptoms of rhinorrhea dren and adults; and the hyper-responsiveness of allergic showed a dose-related trend following methacholine adult subjects to challenge with these substances extends challenge. to allergic children. Of the three mediators used, howThe non-significant effect of bradykinin challenge in ever, histamine appears to be the most preferable. It is not only safe, but can be used to differentiate allergic from this study was probably associated with the rather low nonallergic subjects. The responses most useful to look at doses employed. In one comparative study using adults, to differentiate allergic from non allergic individuals are bradykinin doses of 5 mg were required to provoke significant nasal secretion production, nasal obstruction and secretion weight and symptoms of congestion and rhinorsymptoms of rhinorrhea and congestion.23 However, that rhea. The challenge protocol employed was safe and may OceanSide Publications system to document the effects of other indose was also associated with sinus pain and pressure and offer a model therefore was not considered for inclusion in this study of flammatory substances in children and to evaluate potenIP: 152.118.148.218 children. In contrast, doses of methacholine as high as 10 tial anti-mediator therapies. mg did not produce appreciable symptoms or measurable pathophysiologies in the adult study. Thus, it is doubtful REFERENCES that increasing the dose of methacholine beyond that used 1. Skoner DP, Doyle WJ, Fireman P. 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McLean JA, Mathews KP, Solomon WR, et al. Effect of histamine and methacholine on nasal airway resistance in atopic and nonatopic subjects. J Allergy Clin ImmunoI1977;59: 165-170. van Wijk RG, Dieges PH. Comparison of nasal responsiveness to histamine, methacholine and phentolamine in allergic rhinitis patients and controls. Clin Allergy 1987;17:563-170. Mullins RJ, Olson LG, Sutherland DC. Nasal histamine challenges in symptomatic allergic rhinitis. J Allergy Clin Immunol 1989;83:955-959. Walker SB, Shapiro GG, Bierman CW, et al. Induction of eustachian tube dysfunction with histamine nasal provocation. J Allergy Clin ImmunoI1985;76:158-162. Tomonaga K, Kurono Y, Mogi G. The role of nasal allergy in otitis media with effusion. Acta OtolaryngoI1988;(S458):41-47. Corrado 01, Gould CA, Kassab lY, et al. Nasal response of rhinitic and non-rhinitic subjects to histamine and methacholine: a comparative study. Thorax 1986;4] :863-868. Doyle WI, Boehm S, Skoner DP. Physiologic responses to intranasal dose-response challenges with histamine, methacholine, bradykinin and prostaglandin in adult volunteers with and without nasal allergy. J Allergy Clin ImmunoI1990;86:924-935. Ingall M, Glaser J, Meltzer RS, et al. Allergic rhinitis in early infancy: Review of the literature and report of a case in a newborn. Pediatrics 1965;35:108. Nagy GW, Settipane GA. Bronchial asthma, allergic rhinitis and allergy skin tests among college students. I Allergy 1969; 44:323. 0

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