No evidence for parental age effects on offspring leukocyte telomere length in free-living Soay sheep.

In humans, the effect of paternal age at conception (PAC) on offspring leukocyte telomere length (LTL) is well established, with older fathers thought to pass on longer telomeres to their offspring in their sperm. Few studies have looked for PAC effects in other species, but it has been hypothesised that the effect will be exacerbated in polygamous species with higher levels of sperm competition and production. We test for maternal (MAC) and paternal age at conception effects on offspring LTL in Soay sheep, a primitive breed experiencing strong sperm competition. We use qPCR to measure relative telomere length in 389 blood samples (n = 318 individuals) collected from an unmanaged population of sheep on St Kilda, where individual age and parentage are known. We find no evidence that either MAC or PAC are associated with LTL in offspring across the age range, or when considering only young lambs (n = 164). This is the first study to test for parental age effects on offspring LTL in a wild mammal population, and the results contrast with the findings of numerous human studies that find a PAC effect, as well as predictions of a stronger PAC effect in polygamous species.

Birth of offspring after artificial insemination of heifers with frozen-thawed, sex-sorted, re-frozen-thawed bull sperm.

Two field trials were conducted to determine the fertilising capacity of (i) frozen-thawed, sex-sorted re-frozen-thawed (FSF) dairy bull sperm inseminated close to the time of ovulation, (ii) FSF sperm following large dose insemination, and frozen-thawed, sex-sorted (FS) sperm inseminated within 12h after sorting. In Trial 1, 24 heifers in synchronised oestrus were observed for standing heat over a 30-min period once every 3h. Upon observation of standing heat, the size of the pre-ovulatory follicle was tracked by ultrasound every 6h until ovulation was judged to be imminent. Heifers were inseminated with 4 x 10(6) X-bearing FSF or Control sperm within 6h of ovulation. Ovaries were scanned 6h after AI to ensure ovulation had occurred. All 24 heifers displayed standing oestrus and 20 of these subsequently ovulated. The mean length of standing oestrus was 16.8+/-0.4h and ovulation occurred 27.6+/-1.1h after the onset of standing heat from a pre-ovulatory follicle with a diameter of 16.1+/-0.3mm. All 12 heifers that received FSF sperm returned to oestrus<26d after AI. Of 8 heifers that received Control sperm, 6 (75%) were confirmed pregnant by ultrasound 7 wk after AI, confirming that the method of AI and herd fertility were sound. In Trial 2 the number of sperm inseminated and the effect of eliminating the post-sort cryopreservation step were investigated. Heifers (n=21) were synchronised for oestrus, and inseminated 24h after the onset of standing oestrus with 10 x 10(6) X-bearing FSF, 4 x 10(6) X-bearing FS, or 10 x 10(6) non-sorted frozen-thawed (Control) sperm. Heifers were observed for return to oestrus from 21d, and diagnosed for pregnancy 7 wk after AI. Of the 7 heifers that received FSF sperm, one was confirmed pregnant (14.3%) and delivered a female calf. Four heifers inseminated with control sperm became pregnant and calved, but no pregnancies were obtained using FS sperm. The birth of a calf following AI with FSF sperm demonstrates the potential of sorting from frozen-thawed semen, and with further work, may be a promising technique that will give producers access to sexed sperm from a greater range of bulls.

Pregnancy loss in heifers after artificial insemination with frozen-thawed, sex-sorted, re-frozen-thawed dairy bull sperm.

The use of sex-sorted sperm by the dairy industry is often limited by the geographical distance between potential sires and the sex-sorting facility. One method that may be used to overcome this limitation is sex-sorting sperm that have been previously frozen, or transported to the sorting facility as cooled liquid semen. In this study the in vivo fertility of frozen-thawed, sex-sorted, re-frozen-thawed (FSF) and cooled, sex-sorted, frozen-thawed (CSF) bull sperm was determined after artificial insemination (AI) of Holstein heifers. Semen from two bulls was frozen in straws, or transported to the sorting facility in an egg yolk diluent at 5 degrees C over 24h. Thawed or re-warmed semen was processed through a PureSperm density gradient, and sperm were sorted for sex and frozen (2 or 4 x 10(6)sperm/straw). Synchronised heifers (n=183) were inseminated with either non-sorted control sperm (Control; 20 x 10(6) dose) or with FSF or CSF 'X' sperm (2 or 4 x 10(6)/dose). Pregnancy rates (detected at 7-9 weeks) after AI with control sperm were higher than with FSF or CSF sperm (57.4 vs. 4.1 and 7.3% respectively; p<0.001). There was a significant difference between bulls (Bull 1: Control 63.0%, FSF 8.6%, CSF 10.0%; Bull 2: Control 45.5%, FSF 0%, CSF 4.8%; p=0.001). Five out of six (83.3%) pregnancies produced with sexed sperm were lost after pregnancy diagnosis. The exception was one heifer inseminated with CSF sperm (2 million sperm dose), which produced a heifer calf. In the non-sorted control group, three pregnancies were lost (8.3%) and three stillbirths occurred (8.3%). The low fertility and high rate of pregnancy loss in the sexed groups, in addition to environmental influences, may be attributed to impaired sperm function caused by sex-sorting and re-freezing, leading to poor embryo quality or altered gene expression. More precise timing of insemination and higher sperm doses might improve the fertility of FSF sperm. Moreover, the in vitro function of double-frozen sexed compared with non-sorted sperm requires further investigation.

Embryo production after in vitro fertilization with frozen-thawed, sex-sorted, re-frozen-thawed bull sperm.

The objective of this study was to determine the in vitro fertilizing capacity of bull sperm derived from fresh or frozen samples and subjected to sex sorting and re-cryopreservation. Four sperm types were assessed for their ability to fertilize and sustain early embryo development in vitro. Semen from three Bos taurus bulls of different breeds (Jersey, Holstein and Simmental) was collected and either sorted immediately and then frozen (SF) or frozen for later sorting. Frozen sperm destined for sorting were thawed, sex-sorted, and re-frozen (FSF) or thawed, sex-sorted (FS), and used immediately for in vitro fertilization (IVF). Frozen-thawed nonsorted semen from the same ejaculate was used as a control. Oocytes from donor cows were aspirated via ovum pick-up and matured in vitro prior to IVF and culture. On average, 19.0+/-1.7 (mean+/-SEM) oocytes were aspirated per donor cow, of which 74.4+/-2.2% were selected for maturation. The proportion of cleaved embryos (Day 3) did not differ between sperm groups (P=0.91). Likewise, IVF with FSF sperm resulted in similar Day 7 blastocyst rates (as a percentage of total oocytes) as those of control, SF, and FS sperm (FSF, 34.5+/-4.7; control, 32.2+/-4.6; SF, 35.9+/-4.8; and FS, 26.9+/-4.1%; P=0.23). These encouraging results show that frozen-thawed sex-sorted sperm may be re-frozen and used for in vitro embryo production with similar blastocyst production as that of nonsorted frozen-thawed and sex-sorted frozen-thawed sperm.

In vitro characteristics of frozen-thawed, sex-sorted bull sperm after refreezing or incubation at 15 or 37 degrees C.

The objective was to determine the in vitro characteristics of frozen-thawed dairy bull sperm after sex-sorting and refreezing and thawing (0, 2, and 4h post-thaw; 37 degrees C) or post-sort incubation at 15 or 37 degrees C for 30 and 24h, respectively. These sperm were compared with nonsorted frozen-thawed sperm (control) and with nonsorted sperm undergoing two cryopreservation procedures (FF; 0, 2, and 4h). Frozen-thawed sex-sorted (FS) sperm maintained at 15 or 37 degrees C had higher (P<0.001) progressive motility (PM), velocity, mitochondrial function, viability, and acrosome integrity than that of control sperm but similar total motility at 0 and 2h of incubation. Frozen-thawed sex-sorted sperm incubated at 15 degrees C maintained high levels of motility (66.5+/-1.6%) and viability/acrosome integrity (64.9+/-1.2%) at 24h incubation and, after rewarming and further 6h incubation at 37 degrees C, acceptable levels of motility (35.8+/-1.6%) and viability/acrosome integrity (51.2+/-1.2%) were maintained. Frozen-thawed sex-sorted sperm maintained at 37 degrees C had lower levels of motility, integrity, mitochondrial respiration, and velocity from 4h of incubation onward than that of those incubated at 15 degrees C. However, when frozen-thawed sex-sorted sperm were refrozen (FSF), motility and velocity were depressed at all hours compared with levels exhibited by control sperm, but membrane viability/acrosome integrity and mitochondrial respiration were similar at 0 and 2h post-thaw. Acrosome integrity of sperm in all groups undergoing sorting was exceptionally high at 0h (> or =90%), even after re-cryopreservation and 4h of incubation (77.5+/-1.3%). Double frozen-thawed nonsorted sperm (FF) had similar motility to FSF sperm at 0 and 2h post-thaw but at all time points had the lowest (P<0.001) levels of acrosome intact/viable sperm and mitochondrial respiration and the lowest velocity at 0 h. In conclusion, whereas sex-sorting improved the functionality of frozen-thawed sperm, refreezing depressed motility, viability, and velocity but not acrosome integrity after extended incubation compared with that of control sperm. Furthermore, frozen-thawed, sex-sorted sperm may be stored for transport at 15 degrees C for up 24h without detrimental effects on in vitro sperm characteristics.

Sperm sexing in sheep and cattle: the exception and the rule.

Flow cytometric sorting for the preselection of sex has progressed considerably in the 20 years since its inception. This technique has allowed the production of pre-sexed offspring in a multitude of species and become a commercial success in cattle around the world. However, due to the stress inherent to the sex-sorting process, sex-sorted spermatozoa are widely recognized as functionally compromised in terms of their fertilizing lifespan within the female reproductive tract as a result of reduced motility and viability and changed functional state. These characteristics, when compared to non-sorted controls, are manifest in vivo as lower fertility. However, improvements to the technology and a greater understanding of its biological impact have facilitated recent developments in sheep, showing sex-sorting is capable of selecting a functionally superior population in terms of both in vitro and in vivo function. These results are reviewed in the context of recent developments in other species and the reasons for success after artificial insemination with sex-sorted ram spermatozoa are discussed.

Development of procedures for sex-sorting frozen-thawed bovine spermatozoa.

Dairy bull sperm may be sex-sorted, frozen and used to artificially inseminate heifers with acceptable fertility if the herd is well-managed. One drawback to the technology is that donor bulls must be located within a short distance of the sorting facility in order to collect semen, which limits the number of bulls from which sorted sperm are available. A successful method used to overcome this limitation in sheep is sex-sorting from frozen-thawed semen and refreezing for artificial insemination. This technique is attractive to the dairy industry, and therefore a series of three experiments was designed to investigate the optimal methods to prepare, sex-sort and re-freeze frozen-thawed bovine sperm. Sperm were prepared for sorting by density gradient separation in either PureSperm or BoviPure, followed by staining in one of three diluents (Androhep, Bovine Sheath Fluid + 0.3% BSA or TALP buffer). Sperm were sorted and collected into Test yolk buffer, and frozen in an extender containing 0, 0.25, 0.375 or 0.5% Equex STM Paste. Frozen-thawed sperm were better orientated (p = 0.006) and had fewer damaged membranes (8.7 +/- 0.6% vs 19.5 +/- 2.4%; p = 0.003) after centrifugation in PureSperm rather than BoviPure gradients. Sperm orientation (p < 0.05) and motility (69.9 +/- 3.0 vs 55.6 +/- 4.0; p < 0.001) were highest after staining in Androhep rather than in TALP buffer. Sperm were more motile (58.2 +/- 4.7 vs 38.7 +/- 3.5; p < 0.001) and had better acrosome integrity (74.3 +/- 2.9 vs 66.8 +/- 2.0; p < 0.001) after freezing in an extender containing 0.375% Equex STM Paste than in extender without Equex. Hence, a protocol has been developed to allow frozen-thawed bull sperm to be sex-sorted with high resolution between the sexes, then re-frozen and thawed with retention of motility and acrosome integrity.

Association analysis of the chromosome 4p-located G protein-coupled receptor 78 (GPR78) gene in bipolar affective disorder and schizophrenia.

The orphan G protein-coupled receptor 78 (GPR78) gene lies within a region of chromosome 4p where we have previously shown linkage to bipolar affective disorder (BPAD) in a large Scottish family. GPR78 was screened for single-nucleotide polymorphisms (SNPs) and a linkage disequilibrium map was constructed. Six tagging SNPs were selected and tested for association on a sample of 377 BPAD, 392 schizophrenia (SCZ) and 470 control individuals. Using standard chi(2) statistics and a backwards logistic regression approach to adjust for the effect of sex, SNP rs1282, located approximately 3 kb upstream of the coding region, was identified as a potentially important variant in SCZ (chi(2) P=0.044; LRT P=0.065). When the analysis was restricted to females, the strength of association increased to an uncorrected allele P-value of 0.015 (odds ratios (OR)=1.688, 95% confidence intervals (CI): 1.104-2.581) and uncorrected genotype P-value of 0.015 (OR=5.991, 95% CI: 1.545-23.232). Under the recessive model, the genotype P-value improved further to 0.005 (OR=5.618, 95% CI: 1.460-21.617) and remained significant after correcting for multiple testing (P=0.017). No single-marker association was detected in the SCZ males, in the BPAD individuals or with any other SNP. Haplotype analysis of the case-control samples revealed several global and individual haplotypes, with P-values <0.05, all but one of which contained SNP rs1282. After correcting for multiple testing, two haplotypes remained significant in both the female BPAD individuals (P=0.038 and 0.032) and in the full sample of affected female individuals (P=0.044 and 0.033). Our results provide preliminary evidence for the involvement of GPR78 in susceptibility to BPAD and SCZ in the Scottish population.

Effects of cyclosporin A administered into the airways against antigen-induced airway inflammation and hyperreactivity in the rat.

The immunosuppressant cyclosporin A given orally has anti-asthma properties but carries an undesirable risk of systemic effects. We administered cyclosporin A to Brown Norway rats either orally (p.o.) or topically by intratracheal (i.t.) instillation into the airways before inhaled antigen. Cyclosporin A suppressed the antigen-induced accumulation of activated (CD25+) CD4+ T lymphocytes and eosinophils in the lung, interleukin-5 mRNA expression in lung tissue and airway hyperreactivity. Intratracheal cyclosporin A suppressed cell accumulation at a 10-fold lower dose than that required orally. Minimum effective doses were 3 mg x kg(-1) i.t. and 30 mg x kg(-1) p.o. Intratracheal administration reduced the plasma concentration and systemic exposure compared with an equieffective oral dose, but the reduction (4-5-fold) was not as large as anticipated. Our data suggests that although topical administration to asthmatics would provide some potential for an improved safety margin, it may not offer any major advantage over existing oral therapy. However, the data clearly demonstrate that a novel immunosuppressant with similar anti-inflammatory properties but reduced potential for systemic effects would offer an attractive therapy for severe asthma.

RPR 106541, a novel, airways-selective glucocorticoid: effects against antigen-induced CD4+ T lymphocyte accumulation and cytokine gene expression in the Brown Norway rat lung.

1. The effects of a novel 17-thiosteroid, RPR 106541, were investigated in a rat model of allergic airway inflammation. 2. In sensitized Brown Norway rats, challenge with inhaled antigen (ovalbumin) caused an influx of eosinophils and neutrophils into the lung tissue and airway lumen. In the lung tissue there was also an accumulation of CD4+ T lymphocytes and increased expression of mRNA for interleukin-4 (IL-4) and IL-5, but not interferon-gamma (IFN-gamma). These findings are consistent with an eosinophilia orchestrated by activated Th2-type cells. 3. RPR 106541 (10-300 microg kg[-1]), administered by intratracheal instillation into the airways 24 h and 1 h before antigen challenge, dose-dependently inhibited cell influx into the airway lumen. RPR 106541 (100 microg kg[-1]) caused a significant (P<0.01) (98%) inhibition of eosinophil influx and a significant (P<0.01) (100%) inhibition of neutrophil influx. RPR 106541 was approximately 7 times and 4 times more potent than budesonide and fluticasone propionate, respectively. 4. When tested at a single dose (300 microg kg[-1]), RPR 106541 and fluticasone each caused a significant (P<0.01) (100%) inhibition of CD4+ T cell accumulation in lung tissue. Budesonide (300 microg kg[-1]) had no significant effect. RPR 106541 and fluticasone (300 microg kg[-1]), but not budesonide (300 microg kg[-1]), significantly (P<0.05) inhibited the expression within lung tissue of mRNA for IL-4. RPR 106541 (300 microg kg[-1]) also significantly (P<0.05) inhibited expression of mRNA for IL-5. 5. The high topical potency of RPR 106541 in this model, which mimics important aspects of airway inflammation in human allergic asthmatics, suggests that this glucocorticoid may be useful in the treatment of bronchial asthma.

IgE production, antigen-induced airway inflammation and airway hyperreactivity in the brown Norway rat: the effects of ricin.

Ricin has been shown to enhance IgE production in the rat, probably through inhibition of suppressor T lymphocytes. We have studied further the effects of ricin on IgE titre and have determined its effects on antigen-induced airway inflammation and hyperreactivity in the Brown Norway rat. Immunization with ovalbumin (1-100 micrograms, intraperitoneally) produced dose-related increases in serum antigen-specific IgE titre. Ricin augmented the total IgE titre and caused about a 10-fold increase in the peak antigen-specific IgE titre. In sensitized animals, antigen challenge (three times with aerosolized ovalbumin every second day) caused a significant influx of eosinophils and neutrophils and significant airway hyperreactivity 24 hr after the third challenge. In sensitized animals that had also received ricin, the eosinophil and neutrophil influx was further significantly potentiated and a significant influx of lymphocytes also occurred. Thus, there was a relationship between the degree of sensitization and the magnitude of the inflammatory response. However, the enhanced airway inflammation in ricin-treated animals was not accompanied by a further enhancement of airway hyperreactivity. The present study demonstrates that ricin enhances IgE production and augments an antigen-induced inflammatory pathology but does not potentiate antigen-induced airway hyperreactivity.

Ricin increases IgE levels and airway inflammation but not hyperresponsiveness in the rat.

The effect of ricin treatment on IgE levels and airway inflammation has been examined in ovalbumin (OA)-sensitized rats. Ricin augmented the total IgE titre and caused about a 10-fold increase in the antigen-specific IgE titre. Repeat antigen challenges (every second day) with an OA aerosol increased the number of eosinophils and neutrophils in bronchoalveolar lavage fluid and produced airway hyperresponsiveness (AHR) 24 h after the third challenge. Ricin pretreatment caused a significantly larger influx of eosinophils, neutrophils and lymphocytes into lavage fluid, but there was no further increase in AHR. The present study demonstrates that ricin augments IgE production and the inflammatory response, but that AHR is unrelated to the number of infiltrating inflammatory cells.

Anti-inflammatory and bronchodilator properties of RP 73401, a novel and selective phosphodiesterase type IV inhibitor.

1. We have investigated the effects of RP 73401, a novel, potent and highly selective cyclic nucleotide phosphodiesterase (PDE) type IV inhibitor, in guinea-pig and rat models of bronchoconstriction and allergic inflammation. In some models, the effects of RP 73401 have been compared with those of the standard PDE type IV inhibitor, rolipram. 2. RP 73401 (0.4-400 micrograms kg-1, intratracheally (i.t.) on lactose) inhibited antigen-induced bronchospasm in previously sensitized conscious guinea-pigs (ID50: 7 +/- 1 micrograms kg-1) and in anaesthetized rats (ID50: 100 +/- 25 micrograms kg-1). Rolipram inhibited the antigen-induced bronchospasm in guinea-pigs with an ID50 of 5 +/- 1 micrograms kg-1. In guinea-pig bronchoalveolar lavage (BAL) fluid, total inflammatory cell and eosinophil numbers were reduced by RP 73401 (ID50s: 3.9 +/- 0.8 micrograms kg-1 and 3.2 +/- 0.7 micrograms kg-1, respectively). In the rat, inflammatory cell numbers are less affected. Only the highest dose of RP 73401 (400 micrograms kg-1) significantly inhibited eosinophil influx (41 +/- 16% inhibition). 3. RP 73401 (0.02-100 micrograms kg-1, i.v.) inhibited PAF-induced bronchial hyperreactivity to bombesin in the anaesthetized guinea-pig (ID50: 0.09 +/- 0.03 micrograms kg-1) and inhibited (0.4-40 micrograms kg-1, i.t.) histamine-induced airway microvascular leakage in the anaesthetized guinea-pig by approximately 60% at all doses. 4. RP 73401 relaxed guinea-pig isolated trachea under basal tone (EC50: 9 nM) and when precontracted with histamine (IC50: 2 nM), methacholine (IC50: 29 nM) or leukotriene D4 (LTD4, IC50: 4 nM). 5. RP 73401 (0.4-100 microg kg-1, i.t.) inhibited bronchospasm induced by histamine (ID.%: 34 +/- 6 microg kg-1), methacholine (ID50: 66 +/- 12 pg kg-1) and LTD4 (ID50: <4 microg kg-1) in the anaesthetized guinea pig.Against these same bronchoconstrictors, rolipram (i.t.) had ID5o values of 44 +/- 4, 72 +/- 18 and<4 pg kg- respectively. RP 73401 (4 and 40 pg kg-, i.t.) increased the magnitude and duration of bronchodilatation produced by salbutamol in the anaesthetized guinea-pig. At doses producing significant bronchodilatation, RP 73401 was without effect on heart rate or blood pressure in the anaesthetized guinea-pig. RP 73401 (0.01 -0.25 mg kg-1, i.v.) did not affect heart rate and produced only a small fall in blood pressure in the anaesthetized rat.6. These data demonstrate that RP 73401 and rolipram inhibit antigen- and mediator-induced bronchospasmin guinea-pigs with the same potency. Furthermore, RP 73401 administered directly into the airways, protects against allergic airway inflammation. These results indicate the importance of PDE IV in regulating smooth muscle and inflammatory cell activity. At doses suppressing the inflammatory response in the lung, RP 73401 had little effect in the cardiovascular system. RP 73401 may have a role as a bronchodilator and, more importantly, as a prophylactic anti-inflammatory agent in the treatment of asthma.

RP 58802B, a long-acting beta 2-adrenoceptor agonist: assessment of antiasthma activity in the guinea-pig in vivo.

We have examined the protective actions of RP 58802B, a novel beta 2-adrenoceptor agonist, administered by the inhaled and oral routes in the anaesthetized and conscious guinea-pig against bronchospasm induced by histamine or antigen (ovalbumin). We have also examined the effects of RP 58802B on airway reactivity and inflammatory cell infiltration in platelet-activating factor (PAF) (aerosol)-induced bronchial hyperreactivity and on PAF (tracheal instillation)-induced microvascular leakage in the guinea-pig. Nebulized RP 58802B produced a rapid onset and long lasting inhibition of histamine-induced bronchospasm in the anaesthetized guinea-pig (EC50 = 3.2 +/- 0.9 micrograms/ml; duration greater than 90 min). Given orally, RP 58802B (5 mg/kg, 60 min before challenge) produced a greater than three-fold shift to the right of the dose-response curve and depressed the maximum response to histamine by 39 +/- 11%. Increasing the concentration to 25 mg/kg had no futher effect. Similar protection was still seen 4 h after oral dosing. In conscious guinea-pigs, RP 58802B (5 or 25 mg/kg, p.o. 60 min before challenge) significantly attenuated antigen-induced dyspnoea with the time to severe dyspnoea increasing from 170 +/- 32 to 325 +/- 32 s at the higher dose of drug. RP 58802B (10 or 25 mg/kg, p.o. 60 min before exposure to PAF) prevented the development of bronchial hyperreactivity. Although PAF-induced bronchial hyperreactivity was not accompanied by an increase in the number of pulmonary eosinophils, RP 58802B (25 mg/kg p.o.) reduced the numbers of eosinophils recovered by lavage. RP 58802B (10 mg/kg p.o.) significantly inhibited PAF-induced microvascular leakage into guinea-pig lung. These data suggest that RP 58802B, in addition to being a potent and long acting bronchodilator, may have a prophylactic role in preventing bronchial hyperreactivity and in reducing plasma exudation into the lungs.

Techniques for drug delivery to the airways, and the assessment of lung function in animal models.

A common approach to understanding the mechanisms underlying clinical asthma and in new drug development is to mimic the disease in animal models. When developing animal models of pulmonary diseases, such as asthma, the experimentally induced disease may be characterized in terms of pathophysiological changes induced (e.g., inflammation, smooth muscle contraction) or by the indices of lung function that are effected by such changes. Similarly, the effects of drugs can be assessed in terms of the reversal of disease- or mediator-induced changes in lung function. Small animals, such as the guinea pig and rat, are commonly used for the assessment of lung function in models of pulmonary diseases, such as asthma, and to evaluate the effects of drugs. A variety of techniques, differing in their level of sophistication, has been developed to measure parameters of lung function in small laboratory animals. Simple techniques involve the visual assessment of the response of a conscious animal to bronchoconstriction induced by an inhaled spasmogen or antigen. This technique is rapid but gives results that are difficult to interpret in physiological terms. Bronchospasm can be better assessed in anesthetized, mechanically ventilated animals by recording bronchial tone as changes in either 1) ventilation circuit pressure or 2) air overflow as the lungs are inflated. These techniques are widely used but because they require surgical intervention they are not suited to long-term or repeat studies. In addition, they give only a limited indication of the physiological changes that affect airway caliber. To improve the models available, researchers have subsequently developed techniques that use the same physiological principles as some of the tests applied to the assessment of lung function in humans. These techniques allow the measurement of parameters of respiratory mechanics, such as lung compliance and airway resistance, that determine the relationship between pulmonary pressure changes and air flow into and out of the lungs. Continued development has resulted in models that use nonsurgical plethysmographic techniques. These allow the long-term or repeated measurement of lung function in conscious animals under minimal restraint. In the treatment of asthma, inhalation is the preferred route of administration of a drug as it allows rapid drug delivery to the site of action. Systemic effects are reduced, and the therapeutic dose is minimized. Drugs are generally inhaled as either nebulized liquids or dry-powder formulations. Because drug inhalation requires patient cooperation, techniques have been modified to allow drug delivery to the airways of experimental animals.(ABSTRACT TRUNCATED AT 400 WORDS)

RP 59227, a novel PAF receptor antagonist: effects in guinea pig models of airway hyperreactivity.

The PAF antagonists RP 59227 and WEB 2086 (100 micrograms.kg-1 i.v., 10 min prior to platelet-activating factor (PAF) infusion) abolished or reduced (P less than 0.05) hyperreactivity to bombesin measured at 1 h. Similarly, RP 59227 and WEB 2086 (10 mg.kg-1 p.o., 1 h prior to PAF aerosol) abolished or reduced (P less than 0.01) hyperreactivity to bombesin measured at 24 h. Lower concentrations of RP 59227 and WEB 2086 (3 mg.kg-1 p.o.) were without effect. RP 59227 or WEB 2086 (3 or 10 mg.kg-1 p.o., 1 h prior to antigen aerosol) did not protect against antigen-induced hyperreactivity to histamine measured at 24 h. Antigen-(but not PAF)-induced hyperreactivity was accompanied by an increase in total cell number and, specifically, eosinophil number in bronchoalveolar lavage fluid. The PAF antagonists did not affect BALF cell populations. It is concluded that RP 59227 and WEB 2086 are potent PAF antagonists which inhibit PAF-but not antigen-induced airway hyperreactivity. These data suggest that endogenous PAF may not be involved in antigen-induced hyperreactivity in the guinea pig.

A novel technique for the administration of bronchodilator drugs formulated as dry powders to the anaesthetized guinea pig.

A technique is described for the administration of dry powder formulations of bronchodilator drugs to the anaesthetized guinea pig. The technique has been evaluated by comparing the efficacy of several bronchodilator drugs administered either as dry powders coformulated with a lactose carrier or as nebulized solutions. In each case, the dry powder formulation had comparable bronchodilator activity and duration of action to an equivalent dose administered as a nebulized solution. This technique offers a simple and inexpensive method for the rapid screening of bronchodilator drugs without the problems associated with aerosol administration.