Perspectives on the formation of an interdisciplinary research team.

As research funding becomes more competitive, it will be imperative for researchers to break the mentality of a single laboratory/single research focus and develop an interdisciplinary research team aimed at addressing real world challenges. Members of this team may be at the same institution, may be found regionally, or may be international. However, all must share the same passion for a topic that is bigger than any individual's research focus. Moreover, special consideration should be given to the professional development issues of junior faculty participating in interdisciplinary research teams. While participation may be "humbling" at times, the sheer volume of research progress that may be achieved through interdisciplinary collaboration, even in light of a short supply of grant dollars, is remarkable.

Symptoms of chronic fatigue syndrome/myalgic encephalopathy are not determined by activity pacing when measured by the chronic pain coping inventory.

OBJECTIVES: Chronic fatigue syndrome/myalgic encephalopathy (CFS/ME) is a chronic illness which can cause significant fatigue, pain and disability. Activity pacing is frequently advocated as a beneficial coping strategy, however, it is unclear whether pacing is significantly associated with symptoms in people with CFS/ME. The first aim of this study was therefore to explore the cross-sectional associations between pacing and levels of pain, disability and fatigue. The second aim was to explore whether changes in activity pacing following participation in a symptom management programme were related to changes in clinical outcomes. DESIGN: Cross-sectional study exploring the relationships between pacing, pain, disability and fatigue (n=114) and pre-post treatment longitudinal study of a cohort of patients participating in a symptom management programme (n=35). SETTING: Out-patient physiotherapy CFS/ME service. PARTICIPANTS: One-hundred and fourteen adult patients with CFS/ME. MAIN OUTCOME MEASURES: Pacing was assessed using the chronic pain coping inventory. Pain was measured using a Numeric Pain Rating Scale, fatigue with the Chalder Fatigue Scale and disability with the Fibromyalgia Impact Questionnaire. RESULTS: No significant associations were observed between activity pacing and levels of pain, disability or fatigue. Likewise, changes in pacing were not significantly associated with changes in pain, disability or fatigue following treatment. CONCLUSIONS: Activity pacing does not appear to be a significant determinant of pain, fatigue or disability in people with CFS/ME when measured with the chronic pain coping index. Consequently, the utility and measurement of pacing require further investigation.

Does adding cognitive-behavioural physiotherapy to exercise improve outcome in patients with chronic neck pain? A randomised controlled trial.

OBJECTIVES: To determine whether adding a physiotherapist-led cognitive-behavioural intervention to an exercise programme improved outcome in patients with chronic neck pain (CNP). DESIGN: Multicentre randomised controlled trial. SETTING: Four outpatient physiotherapy departments. PARTICIPANTS: Fifty-seven patients with CNP. Follow-up data were provided by 39 participants [57% of the progressive neck exercise programme (PNEP) group and 79% of the interactive behavioural modification therapy (IBMT) group]. INTERVENTIONS: Twenty-eight subjects were randomised to the PNEP group and 29 subjects were randomised to the IBMT group. IBMT is underpinned by cognitive-behavioural principles, and aims to modify cognitive risk factors through interactive educational sessions, graded exercise and progressive goal setting. MAIN OUTCOME MEASURES: The main outcome measure was disability, measured by the Northwick Park Questionnaire (NPQ). Secondary outcomes were the Numeric Pain Rating Scale (NPRS), Pain Catastrophising Scale, Tampa Scale for Kinesiophobia (TSK), Chronic Pain Self-efficacy Scale (CPSS) and the Pain Vigilance and Awareness Questionnaire. RESULTS: No significant between-group differences in disability were observed (mean NPQ change: PNEP=-7.2, IBMT=-10.2). However, larger increases in functional self-efficacy (mean CPSS change: PNEP=1.0, IBMT=3.2) and greater reductions in pain intensity (mean NPRS change: PNEP=-1.0, IBMT=-2.2; P<0.05) and pain-related fear (mean TSK change: PNEP=0.2, IBMT=-4.7, P<0.05) were observed with IBMT. Additionally, a significantly greater proportion of participants made clinically meaningful reductions in pain (25% vs 55%, P<0.05) and disability (25% vs 59%, P<0.05) with IBMT. CONCLUSIONS: The primary outcome did not support the use of cognitive-behavioural physiotherapy in all patients with CNP. However, superior outcomes were observed for several secondary measures, and IBMT may offer additional benefit in some patients. CLINICAL TRIAL REGISTRATION NUMBER: ISRCTN27611394.

Identification of cDNA clones that code for protein-tyrosine kinases by screening expression libraries with antibodies against phosphotyrosine.

A phage lambda gt11 human fibroblast cDNA expression library was screened with antibodies against phosphotyrosine to determine the feasibility of this approach as a method for the identification of clones that code for protein-tyrosine kinases. Several antibody positive clones were isolated. One clone also scored positive with degenerate oligonucleotides designed to identify sequences coding for protein-tyrosine kinases. This cDNA clone was partially sequenced and proved to be identical to part of lyn, a recently reported putative protein-tyrosine kinase. A portion of the cDNA was cloned into an inducible plasmid expression vector. Phosphorylation of many bacterial proteins on tyrosine was observed upon addition of inducing agent. The results demonstrate that this method of screening can identify cDNAs that encode active protein-tyrosine kinases and could prove useful for the identification of novel ones.

Neurophysiology of adult male Schistosoma mansoni.

From the studies on the neurophysiology of schistosomes it appears that in spite of the unique hexilaminar arrangement of the tegument's outer membrane, it has biophysical properties not markedly different from those of a variety of other multi-dimensional syncytia. The close electrical coupling of the muscle and tegument must be taken into account when one attempts to define sites and modes of action of drugs which affect motor activity. Agents which disrupt muscle function in the schistosome may exert their action indirectly by way of an effect on the tegumental membrane. The syncytial nature of the musculature and the possibility that longitudinal contraction waves are myogenic suggests that neurotransmitters may simply function as modulators of muscle activity as is the case for many vertebrate visceral muscles. External recordings indicate the presence of a variety of electrically active tissues within the schistosomes. There is no clear correlation of this activity with longitudinal muscle activity or with active membrane responses in this muscle. From this it would appear the bulk of this activity may have its origins in tissues other than the longitudinal muscle such as other muscle groups, nerve trunks, or the peripheral nerve net.

Mechanistic studies in the transcuticular delivery of antiparasitic drugs. II: Ex vivo/in vitro correlation of solute transport by Ascaris suum.

Using live, intact Ascaris suum and a closed perfusion system, the absorption kinetics and tissue distribution of selected radiolabeled permeants were measured to determine the importance of the transcuticular pathway for drug absorption. The data support the conclusions established by previous in vitro transport studies which utilized excised cuticle-hypocuticle tissue preparations. The external surface of A. suum can be breached by drugs and the rate-determining barrier is the lipoidal hypocuticle tissue, provided the permeant is sufficiently small to traverse the aqueous-filled, negatively charged collagen matrix of the cuticle. The ex vivo permeability coefficients of the model permeants for the cuticle-hypocuticle barrier were in good quantitative agreement with the in vitro permeability coefficients. The lipophilic permeants hydrocortisone and p-nitrophenol were preferentially distributed in the gut tissue, whereas the hydrophilic permeant urea was distributed evenly throughout the organism and was extensively metabolized. Ligated and nonligated A. suum showed no significant differences in either uptake kinetics or tissue distribution of the permeants. This indicates that the transcuticular pathway is the major route of drug absorption as compared to oral ingestion.

Changes in glucose metabolism and cyanide sensitivity in Schistosoma mansoni during development.

Schistosoma mansoni was studied by biochemical and electrophysiological techniques to follow the physiological changes occurring during transformation in the mammalian host. Volume conducted electrical potentials and measurement of CO2 evolution indicate that 3 h post-transformational schistosomula are highly sensitive to cyanide. By 24 h after transformation, evolution of CO2 under control conditions is reduced by 77% from 3 h levels, while lactate excretion rises by 84%. Cyanide does not affect the frequency or magnitude of endogenous electrical transients, but does eliminate 83% of the already reduced levels of CO2 evolved in 24 schistosomula. Electrophysiological analyses indicate that the timecourse of metabolic changes in skin- and mechanically transformed schistosomula are similar, and incubation of schistosomula in 200 micrograms ml-1 puromycin does not alter the onset of cyanide insensitivity. The adult parasite evolves a low level of CO2 which is reduced by 88% in the presence of 1 mM cyanide. No significant Pasteur effect is detected, however, and endogenous electrical activity as well as mechanical responses of the adult musculature are unaffected by cyanide exposure. Our results indicate that schistosomula continue to rely on cyanide-sensitive respiratory components for at least 3 h after transformation; by 24 h, however, the parasites are metabolically similar to the adult stage, i.e., they depend on lactate fermentation for most of their energy requirements.

Biophysical transport properties of the cuticle of Ascaris suum.

The transport properties of isolated cuticle from Ascaris suum were studied using standard two-chamber diffusion cells and a number of radiolabeled permeants which varied in molecular size, lipophilicity and electrical charge. The permeability coefficient of the collagen matrix (lipid-extracted cuticle) vs. molecular radius relationship showed the interdependence of molecular size and electrical charge of the permeants with respect to the aqueous pores of the negatively charged matrix. The permeability of neutral solutes decreased monotonically with size. Protonated amines permeated the aqueous pores faster than neutral solutes of comparable size, while the permeation of anions was slower. The average pore size was estimated to be 1.5 nm in radius. A biophysical model which accounted for diffusion of molecules within a fixed electrostatic field of force and for molecular sieving by the pore channels was used in the mechanistic interpretation of the data. The effective permeability coefficient of the non-lipid-extracted cuticle was delineated into the permeability coefficients of the water-filled collagen matrix and the lipoidal component of the cuticle to determine which layer was the rate-controlling barrier. While each solute was capable of penetrating the water-filled collagen matrix, the rate-determining step for the majority of compounds was passive diffusion across the lipid component, which controlled 75-99% of transport. The exception was water, for which transport kinetics was 75% matrix-controlled. In general, permeation across the lipid-filled tissue was more favorable for small lipophilic compounds because of molecular restriction not only in the aqueous pores, but also in the lipid-filled pores.

Mechanisms of microenvironmental pH regulation in the cuticle of Ascaris suum.

The excretion kinetics of various organic acids by Ascaris suum were quantified to determine if the excretion of these metabolic end-products could generate and maintain a microclimate pH within the aqueous compartment of the cuticle. Ligated and nonligated A. suum were incubated in media buffered with 0.25 or 2.5 mM Hepes (initial pH 7.5) or 0.5 or 5 mM glycine (initial pH 3.25). The concentration of organic acids and the pH of the media were followed for 24 h. Several volatile fatty acids, including acetic, 2-methylbutyric, 2-methylvaleric, n-valeric, and n-butyric, were excreted at relatively high rates. Propionic, n-caproic, 2-methylcaproic, tiglic acid, and the non-volatile organic acids, lactic and succinic, were excreted more slowly. The organic acids were excreted at a constant rate and in apparently fixed molar concentration ratios. The accumulation of organic acids was associated with changes in pH of the medium until a limiting constant pH, in the vicinity of the pKa of the volatile fatty acids, was reached. The rate of organic acid excretion was not affected by initial medium pH, buffer capacity, or parasite ligation. The rate of pH change induced by the excretion of organic acids was also insensitive to whether ligated or nonligated A. suum were used, but was dependent on the initial buffer capacity of the medium. These results suggest that A. suum excrete the end-products of carbohydrate metabolism across the cuticle. The presence of organic acids in the aqueous pores of the cuticle creates and maintains a microclimate pH of about 5.0 +/- 0.3. This pH will influence the transport properties of weak acids and bases and should be considered in the design of delivery systems for anthelmintics.

Transport of model peptides across Ascaris suum cuticle.

Several FMRFamide-related peptides (FaRPs) found in nematodes exert potent excitatory or inhibitory effects on the somatic musculature of Ascaris suum and other nematode species when injected into the pseudocoelom or applied directly to isolated neuromuscular preparations. These peptides, however, generally fail to induce detectable effects on the neuromusculature when applied externally to intact nematodes. The apparent lack of activity for these peptides when administered externally in whole-organism assays is likely a function of both absorption and metabolism. To delineate the factors that govern transport of peptides across the cuticle/hypodermis complex of nematodes, we measured the rates of absorption of a series of structurally related model peptides using isolated cuticle/hypodermis segments from A. suum and two-chamber diffusion cells. [14C]-Labeled peptides were prepared from D-phenylalanine, with the amide nitrogens sequentially methylated to give AcfNH2, Acf3NH2, Acf(NMef)2NH2, and Ac(NMef)3NHMe. These model peptides were designed to allow systematic analysis of the influence of peptide size, hydrogen bonding and lipophilicity on transport. Results of these studies show that, within this series, permeability across the cuticle increases with addition of each methyl group. The permeability coefficient of Ac(NMef)3NHMe, with four methyl groups, was 10-fold greater than that of the smaller peptide, AcfNH2, even though both peptides contain five hydrogen bonds. When compared with vertebrate membranes, transport of the model peptides across A. suum cuticle was about 10-fold slower. A biophysical model for transcuticular transport of peptides predicted that nematode FaRPs, which are larger, less methylated and less lipophilic than the model peptides tested, would not be absorbed across the cuticle of nematodes. This prediction was confirmed for the excitatory FaRP, AF2 (KHEYLRFamide), which did not diffuse across the cuticle/hypodermis complex, but diffused rapidly across lipid-extracted cuticle preparations.

Structural characterisation and pharmacology of KHEYLRFamide (AF2) and KSAYMRFamide (PF3/AF8) from Haemonchus contortus.

The FMRFamide-related peptides (FaRPs), KHEYLRFamide (AF2) and KSAYMRFamide (PF3) were structurally characterised from the parasitic nematode of sheep, Haemonchus contortus (MH isolate). Both peptides were sequenced in a single gas-phase sequencing run and their structure confirmed by mass spectrometry which identified peptides of 920 Da (C-terminally amidated AF2) and 902/918 Da (C-terminally amidated non-oxidised/oxidised PF3, respectively). AF2 had inhibitory effects on H. contortus muscle and inhibited acetylcholine (ACh, 10 microM)-induced contractions, with a threshold for activity of 1 microM. PF3 induced concentration-dependent contractions of H. contortus (activity threshold, 10 nM) and enhanced ACh contractions. Compared with the MH isolate, an isolate of H. contortus which has reduced sensitivity to cholinergic drugs (Lawes isolate) was less sensitive to the effects of PF3. The concentration-response curves for the cholinergic compounds ACh and levamisole (LEV), and PF3, but not a control, KPNFIRFamide (PF4), showed a statistically similar shift. This study implicates PF3 in the modulation of cholinergic function in H. contortus.

Biophysical model for organic acid excretion in Ascaris suum.

To develop a model for the mechanisms of organic acid excretion in nematodes, we measured the concentrations of volatile fatty acids (VFAs), pH and electrical potentials across hypodermal and muscle membranes and across the composite body wall (consisting of hypodermis, muscle and cuticle) of Ascaris suum using standard chromatographic and microelectrode recording techniques. In incubates containing one parasite in 20 ml modified Ascaris Ringer's solution, the level of combined VFAs excreted into the medium increased linearly for about 18 h, then plateaued at a concentration of 4.2 mM; the medium acidified rapidly to a plateau at about pH 5.0 within 4-6 h. Following 24 h incubations, the concentrations of VFAs in the hypodermis, muscle, and pseudocoelomic compartments were 62.4 +/- 8.1, 62.3 +/- 7.8 and 74.4 +/- 3.2 mM, respectively. The pseudocoelomic fluid was more acidic (pH 6.52 +/- 0.06) than the hypodermis (pH 6.78 +/- 0.03) or muscle (pH 6.77 +/- 0.03). These data and the electrical potentials across hypodermal (-57.9 +/- 6.3 mV) and muscle (-30.3 +/- 0.8 mV) membranes were used to determine the equilibrium concentrations for protonated (HVFA) and anionic (VFA-) forms of the acids across these membranes and across the cuticle. Under these conditions, little transmembrane or transmural excretion of HVFAs is expected to occur in A. suum. However, a 16-27 mV driving force for VFA- excretion exists across hypodermal and muscle membranes, and a larger driving force is predicted to exist for these anions across the cuticle. This driving force could provide potential energy for VFA- excretion through anion channels which exist in muscle and hypodermal membranes of this parasite, or for facilitated transport systems.

Mechanism-based screening: discovery of the next generation of anthelmintics depends upon more basic research.

The therapeutic arsenal for the control of helminth infections contains only a few chemical classes. The development and spread of resistance has eroded the utility of most currently available anthelmintics, at least for some indications, and is a constant threat to further reduce the options for treatment. Discovery and development of novel anthelmintic templates is strategically necessary to preserve the economic and health advantages now gained through chemotherapy. As the costs of development escalate, the question of how best to discover new drugs becomes paramount. Although random screening in infected animals led to the discovery of all currently available anthelmintics, cost constraints and a perception of diminishing returns require new approaches. Taking a cue from drug discovery programmes for human illnesses, we suggest that mechanism-based screening will provide the next generation of anthelmintic molecules. Critical to success in this venture will be the exploitation of the Caenorhabditis elegans genome through bioinformatics and genetic technologies. The greatest obstacle to success in this endeavour is the paucity of information available about the molecular physiology of helminths, making the choice of a discovery target a risky proposition.

The pharmacology of FMRFamide-related neuropeptides in nematodes: new opportunities for rational anthelmintic discovery?

The chemotherapeutic control of helminth parasites is compromised by the limited number of classes of anthelmintic drugs. Discovery of novel anthelmintics is impeded by the lack of novel screening technologies that overcome the difficulties inherent in screens based on whole organism toxicity. The development and implementation of mechanism-based screens for new anthelmintics offers great promise for the revitalization of antiparasitic drug discovery. However, mechanism-based screens must be based on a thorough understanding of the proteins or processes that offer the best chance for selective chemotherapeutic intervention. Basic research on the characterization of nematode FMRFamide-related peptides (FaRPs) has revealed that these peptides are ubiquitously distributed in helminths. Chemical identification of a number of nematode FaRPs has been achieved, and these peptides have potent and profound effects on the nematode neuromuscular system. Physiological processes mediated by nematode FaRPs (and other helminth neuropeptides) offer potential targets for the discovery of novel anthelmintics.

Nematode FMRFamide-related peptide (FaRP)-systems: occurrence, distribution and physiology.

The application of rational (mechanism-based) approaches to anthelmintic discovery requires information about target proteins which are pharmacologically distinguishable from their vertebrate homologs. In helminths, several such targets (e.g., beta-tubulin, ATP-generating enzymes, cholinergic receptors, CI- channels) have been characterized only after the discovery, through empirical screening, of compounds that interfere with their function. From the perspective of anthelmintic discovery, the utility of these targets is diminishing due to the emergence of drug-resistant strains of parasites. This has motivated the search for compounds with novel modes-of-action. Recent basic research in helminth physiology and biochemistry has identified several potential targets for rational anthelmintic discovery, including receptors for FMRFamide-related peptides (FaRPs). To date, over 20 different nematode FaRPs have been identified and these peptides, which are broadly distributed in helminths, have been localized to all of the major neuronal subtypes in nematodes. The FaRPs that have been examined have been found profoundly to affect somatic muscle function in gastrointestinal nematodes. In this respect, complex inhibitory and excitatory actions have been identified for a number of these peptides. Although the transduction pathways for any of these peptides remain to be elucidated, the available evidence indicates that nematode FaRPs have numerous mechanisms of action. The employment of nematode neuropeptide receptors in mechanism-based screens has immense potential in the identification of novel anthelmintics.

The ovijector of Ascaris suum: multiple response types revealed by Caenorhabditis elegans FMRFamide-related peptides.

Caenorhabditis elegans possesses 22 FMRFamide-like peptide (flp) genes predicted to encode 60 different FMRFamide-related peptides with a range of C-terminal signatures. Peptides from five flp genes (1, 6, 8, 9 and 14) are known to modulate the ovijector of Ascaris suum in vitro. This study examines the physiological effects of peptides from the remaining 17 flp genes such that the variety of FMRFamide-related peptide-induced ovijector response types can be delineated. Five categories of response were identified according to the pattern of changes in contractile behaviour and baseline tension. Peptides encoded on 16 flp genes (1, 2, 3, 4, 6, 7, 9, 10, 11, 12, 13, 14, 15, 16, 17 and 20) had qualitatively similar inhibitory (response type 1) actions, with the lowest activity thresholds (1 nM) recorded for peptides with FIRFamide or FLRFamide C-terminal signatures. Peptides encoded on four flp genes (2, 18, 19 and 21), and on the A. suum afp-1 gene, had excitatory actions on the ovijector (response type 2), with PGVLRFamides having the lowest activity threshold (1 nM). An flp-2 peptide (LRGEPIRFamide) induced a transient contraction of the ovijector (activity threshold, 10nM) that was designated response type 3. Response type 4 comprised a transient contraction followed by an extended period of inactivity and was observed with peptides encoded on flp-5 (AGAKFIRFamide, APKPKFIRFamide), flp-8 (KNEFIRFamide) and flp-22 (SPSAKWMRFamide). SPSAKWMRFamide was the most potent peptide tested with an activity threshold of 0.1 nM. A single peptide (AMRNALVRFamide; activity threshold 0.1 microM), encoded on flp-11, induced response type 5, a shortening of the ovijector coupled with an increase in contraction frequency. Although most flp genes encode structurally related peptides that trigger one of the five ovijector response types, flp-2 and flp-11 co-encode FMRFamide-related peptides that induce distinct responses. Within the ovijector of A. suum FaRPs play a complex role involving at least five receptor subtypes or signalling pathways.

Influence of organic acid excretion on cuticle pH and drug absorption by Haemonchus contortus.

To determine if a cuticle microenvironment pH is maintained by adult Haemonchus contortus, organic acid excretion kinetics and absorption kinetics of selected model weak acids and a weak base were measured in incubation media that varied in buffer capacity (0.25-20 mM HEPES or 5 mM glycine) and initial pH (7.5 or 3.5). To evaluate the importance of the cuticle as a pathway for organic acid excretion and drug absorption the pharynx was paralyzed with 1 nM ivermectin. H. contortus changed the media pH from initial values of 7.5 or 3.25 to an asymptotic value of approximately 5.6. The rate of pH change depended on the buffer capacity, but was not affected by chemical ligation with ivermectin. The intrinsic rate of excretion of organic acids (0.045 +/- 0.016 micromol/cm2 x h) was constant during the first 8-12 h of incubation and was independent of initial pH, buffer capacity or ivermectin ligation. The rates of absorption of the model weak acids, benzoic acid and p-nitrophenol, and the model weak base, aniline, were not affected by initial pH, buffer capacity or ivermectin ligation. These results suggest that H. contortus excretes organic acid endproducts of carbohydrate metabolism across its cuticle, and that these acids maintain a microenvironment pH within the water-filled pores of the cuticle that controls the rate of adsorption of weakly acidic or basic drugs.

Effects of KHEYLRFamide and KNEFIRFamide on cyclic adenosine monophosphate levels in Ascaris suum somatic muscle.

KHEYLRF-NH(2) (AF2) is a FMRFamide-related peptide (FaRP) present in parasitic and free-living nematodes. At concentrations as low as 10 pM, AF2 induces a biphasic tension response, consisting of a transient relaxation followed by profound excitation, in neuromuscular strips prepared from Ascaris suum. In the present study, the effects of AF2 on cyclic adenosine monophosphate (cAMP), cyclic guanosine monophosphate (cGMP) and inositol-1,4,5-triphosphate (IP(3)) levels were measured following muscle tension recordings from 2 cm neuromuscular strips prepared from adult A. suum. AF2 induced a concentration- and time-dependent increase in cAMP, beginning at 1 nM; cAMP levels increased by 84-fold following 1 h exposure to 1 microM AF2. cGMP and IP(3) levels were unaffected by AF2 at concentrations

Pharmacology of FMRFamide-related peptides in helminths.

Nervous systems of helminths are highly peptidergic. Species in the phylum Nematoda (roundworms) possess at least 50 FMRFamide-related peptides (FaRPs), with more yet to be identified. To date, few non-FaRP neuropeptides have been identified in these organisms, though evidence suggests that other families are present. FaRPergic systems have important functions in nematode neuromuscular control. In contrast, species in the phylum Platyhelminthes (flatworms) apparently utilize fewer FaRPs than do nematodes; those species examined possess one or two FaRPs. Other neuropeptides, such as neuropeptide F (NPF), play key roles in flatworm physiology. Although progress has been made in the characterization of FaRP pharmacology in helminths, much remains to be learned. Most studies on nematodes have been done with Ascaris suum because of its large size. However, thanks to the Caenorhabditis elegans genome project, we know most about the FaRP complement of this free-living animal. That essentially all C. elegans FaRPs are active on at least one A. suum neuromuscular system argues for conservation of ligand-receptor recognition features among the Nematoda. Structure-activity studies on nematode FaRPs have revealed that structure-activity relationship (SAR) "rules" differ considerably among the FaRPs. Second messenger studies, along with experiments on ionic dependence and anatomical requirements for activity, reveal that FaRPs act through many different mechanisms. Platyhelminth FaRPs are myoexcitatory, and no evidence exists of multiple FaRP receptors in flatworms. Interestingly, there are examples of cross-phylum activity, with some nematode FaRPs being active on flatworm muscle. The extent to which other invertebrate FaRPs show cross-phylum activity remains to be determined. How FaRPergic nerves contribute to the control of behavior in helminths, and are integrated with non-neuropeptidergic systems, also remains to be elucidated.

Isolation and preliminary biological characterization of KPNFIRFamide, a novel FMRFamide-related peptide from the free-living nematode, Panagrellus redivivus.

A novel FMRFamide-related heptapeptide, Lys-Pro-Asn-Phe-Ile-Arg-Phe-NH2 (KPNFIRFamide), was isolated and characterized from acid ethanol extracts of the free-living nematode, Panagrellus redivivus. Whole-worm extracts contained > or = 9 pmol KPNFIRFamide/g wet weight. A synthetic replicate of this peptide induced a rapid relaxation of tone and inhibited spontaneous contractility in isolated innervated and denervated body-wall muscle strips of the parasitic nematode, Ascaris suum. KPNFIRFamide (0.1 nM) induced measurable relaxations in 50% of the muscle preparations examined. Concentrations > or = 0.3 nM induced relaxation in 100% of muscle preparations examined. The relaxation was short-lived at concentrations of peptide > or = 1 microM and displayed a profile typical of receptor desensitization. These data suggest the occurrence of a closely related peptide in A. suum and add further evidence to the concept of primary structural conservation of FaRPs within the nematodes.