ABSTRACT
The interrelationships between physicochemical properties, absorption and potency of 2-desoxoparaherquamide and five analogs, representing a new anthelmintic class, were evaluated in in vitro and in vivo assays. At pH 7.5, rates of drug absorption by the gastrointestinal nematode Haemonchus contortus and jird small intestine, parameterized by the permeability coefficient, P(e), ranged from 1.2-2.4 x 10(-4) cm/min (nematode) to 2.5-5.5 x 10(-3) cm/min (jird). In the jird intestine, absorption was pH-dependent, with P(e) at pH 7.5 being twice that at pH 4.5, reflecting the negative influence of protonation on transport of these weakly basic molecules. Each compound rapidly paralyzed H. contortus during in vitro exposure to therapeutically relevant concentrations (1-10 microm). The kinetics of drug action on motility in vivo mirrored their in vitro effects; motility concentrations were reduced in nematodes collected from jird stomach 3 h following oral drug dosing, by which time > or =50% clearance of the parasites had occurred. The nematode/medium partition coefficient K ranged from 10.1 to 16.1, consistent with the lipophilic nature of the compounds. The time required to reduce motility in vitro by 50% (t50*) and P(e) were used to determine C(n)*, the concentration of drug in the nematode at t50*, as an indicator of intrinsic potency. In the jird, the apparent potencies of the compounds were insensitive to route of administration (i.e. oral = i.v. = i.p. = i.m.) for H. contortus and two other gastrointestinal nematodes, Ostertagia ostertagi and Trichostrongylus colubriformis; topical administration, however, required three to 10-fold higher doses for equivalent efficacy.
Subject(s)
Anthelmintics/pharmacology , Haemonchiasis/veterinary , Haemonchus/drug effects , Indolizines/pharmacology , Sheep Diseases/drug therapy , Spiro Compounds/pharmacology , Absorption , Administration, Oral , Animals , Anthelmintics/administration & dosage , Anthelmintics/pharmacokinetics , Female , Haemonchiasis/drug therapy , Haemonchus/metabolism , Indolizines/administration & dosage , Indolizines/pharmacokinetics , Injections, Intramuscular/veterinary , Injections, Intraperitoneal/veterinary , Injections, Intravenous/veterinary , Parasitic Sensitivity Tests , Random Allocation , Sheep , Sheep Diseases/parasitology , Spiro Compounds/administration & dosage , Spiro Compounds/pharmacokinetics , Trichostrongyloidea/drug effects , Trichostrongyloidea/metabolismABSTRACT
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 =1 microM. AF2-induced stimulation of cAMP was unaffected by removal of the dorsal or ventral nerve cord, even though this form of denervation abolished the excitatory phase of the tension response. The effects of 0.1 and 1 microM AF2 on cAMP were also unaffected by 10 microM SDPNFLRF-NH(2) (PF1, an inhibitory FaRP) and 10 microM PF1022A (an inhibitory cyclodepsipeptide), even though each of these peptides abolished the excitatory phase of the tension response induced by AF2. Within an alanine-scan series of AF2 analogues, only KHAYLRF-NH(2) stimulated cAMP production with equipotency to AF2; the effects of this peptide on muscle tension also mimicked AF2. Another excitatory FaRP present in nematodes, KNEFIRF-NH(2) (AF1), also stimulated cAMP production, but was 100-fold less potent than AF2. The stimulatory effects of AF1 on tension and cAMP levels were blocked by an alanine-substituted analogue of this peptide (Ala(6)-AF1, KNEFIAF-NH(2)), while the stimulatory effects of AF2 on tension and cAMP were not affected by this analogue. AF2 and AF1 increase A. suum somatic muscle cAMP by targeting different receptors. Increases in cAMP stimulated by AF2 can be decoupled from the excitatory response caused by this peptide, and it is not possible to establish a causal linkage between the contractile response elicited by this peptide and its effects on cAMP accumulation.