Post-epizootic microbiome associations across communities of neotropical amphibians.

Microbiome-pathogen interactions are increasingly recognized as an important element of host immunity. While these host-level interactions will have consequences for community disease dynamics, the factors which influence host microbiomes at larger scales are poorly understood. We here describe landscape-scale pathogen-microbiome associations within the context of post-epizootic amphibian chytridiomycosis, a disease caused by the panzootic chytrid fungus Batrachochytrium dendrobatidis. We undertook a survey of Neotropical amphibians across altitudinal gradients in Ecuador ~30 years following the observed amphibian declines and collected skin swab-samples which were metabarcoded using both fungal (ITS-2) and bacterial (r16S) amplicons. The data revealed marked variation in patterns of both B. dendrobatidis infection and microbiome structure that are associated with host life history. Stream breeding amphibians were most likely to be infected with B. dendrobatidis. This increased probability of infection was further associated with increased abundance and diversity of non-Batrachochytrium chytrid fungi in the skin and environmental microbiome. We also show that increased alpha diversity and the relative abundance of fungi are lower in the skin microbiome of adult stream amphibians compared to adult pond-breeding amphibians, an association not seen for bacteria. Finally, stream tadpoles exhibit lower proportions of predicted protective microbial taxa than pond tadpoles, suggesting reduced biotic resistance. Our analyses show that host breeding ecology strongly shapes pathogen-microbiome associations at a landscape scale, a trait that may influence resilience in the face of emerging infectious diseases.

Another frog on the menu: predation of Trachops cirrhosus (Chiroptera: Phyllostomidae) upon Osteocephalus oophagus (Anura: Hylidae)

We report a predation event of the fringe-lipped bat (Trachops cirrhosus) (Chiroptera: Phyllostomidae) upon the slender-legged tree frog (Osteocephalus oophagus) (Anura: Hylidae), in Central Amazonia, Brazil. This observation adds a new prey item to the T. cirrhosus diet and shines some light on the scarce ecological knowledge of both predator and prey species.(AU)

Another frog on the menu: predation of Trachops cirrhosus (Chiroptera: Phyllostomidae) upon Osteocephalus oophagus (Anura: Hylidae)

We report a predation event of the fringe-lipped bat (Trachops cirrhosus) (Chiroptera: Phyllostomidae) upon the slender-legged tree frog (Osteocephalus oophagus) (Anura: Hylidae), in Central Amazonia, Brazil. This observation adds a new prey item to the T. cirrhosus diet and shines some light on the scarce ecological knowledge of both predator and prey species.

Blockade of acetylcholine release at the motor endplate by a polypeptide from the venom of Phoneutria nigriventer.

1. The mechanisms underlying the muscle relaxation effect of a fraction (PF3) isolated from the Phoneutria nigriventer spider venom were assessed on mouse diaphragm and chick biventer cervicis muscle preparations. 2. PF3 (0.25-4 micrograms ml-1) produced a concentration-dependent blockade of the nerve-elicited muscle twitch of the mouse diaphragm (IC50 = 0.8 micrograms ml-1) without affecting the directly induced muscle twitch. In similar preparations, the crude venom (1-10 micrograms ml-1) produced muscle contracture and blocked both the direct and indirectly induced muscle twitches. 3. In the chick biventer cervicis muscle, PF3 (1-5 micrograms ml-1) blocked the nerve stimulated muscle twitch (IC50 = 1.26 micrograms ml-1), but did not alter the postjunctional response to exogenous acetylcholine (ACh, 10 microM-10 mM). 4. PF3 (2-8 micrograms ml-1) reduced the frequency of miniature endplate potentials (m.e.p.ps) recorded intracellularly from the mouse diaphragm muscle fibers by 58 to 64%, and diminished the amplitude of m.e.p.ps by 20 to 40% of control. The relationship between log m.e.p.p. frequency and log [Ca2+]o was shifted rightwards in the presence of 4 micrograms ml-1 PF3. 5. Raising the frequency of m.e.p.ps with high K+ medium or theophylline (3 mM) did not prevent the toxin-induced depression of spontaneous ACh release. 6. The quantal content of e.p.ps (m), determined in cut-diaphragm muscle fibres, was reduced by 53% and 77% of control by 1 and 4 micrograms ml-1 PF3, respectively. At 1 microgram ml-1 the toxin shifted the relationship between log m and log [Ca2+]o towards higher values without apparent change of the slope. 7. E.p.p. trains elicited at 10 to 50 Hz in the presence of PF3 (1 microgram ml-1) exhibited irregular amplitudes and facilitation related to the frequency of nerve stimulation. 8. It is concluded that PF3 blocks neuromuscular transmission by acting prejunctionally and reducing the nerve-evoked transmitter release. The effect was related to a diminished Ca2+ entry into the nerve terminal associated with inhibition of exocytosis.

Prejunctional effect of quaternary derivatives of l-hyoscyamine at the rat neuromuscular junction. A structure-activity relationship study.

1. The effects of phenthonium and related compounds on the spontaneous release of acetylcholine (ACh) were investigated with electrophysiological and radiolabelled techniques to correlate the prejunctional effect with their cholinolytic activities and to determine the structure-activity relationship. 2. Phenthonium and endophen are N-(4-phenyl)-phenacyl derivatives of l-hyoscyamine in "exo" and "endo" conformation, respectively. Tropol is N-(4-phenyl) phenacyl tropan-3-ol whereas ipratropium is 8-isopropyl-noratropine. 3. Only phenthonium increased the frequency of miniature endplate potentials and the resting efflux of spontaneous [3H]-ACh in rat diaphragm muscles. 4. The rank order of the antimuscarinic potency was: ipratropium > atropine > phenthonium = endophen > tropol. The rank order of the antinicotinic activity was: phenthonium = endophen > tropol > atropine > ipratropium. 5. It is concluded that the prejunctional facilitatory effect of phenthonium is associated with the N-phenyl-phenacyl group at "exo" conformation but the effect is unrelated to its cholinolytic properties.

Androgen regulation of the nicotinic acetylcholine receptor-ionic channel in a hormone-dependent skeletal muscle.

The trophic influence of testosterone on the nicotinic acetylcholine receptor-ionic channel (AChR) was studied in the levator ani (LA) muscle of adult male rats (120 days) intact (C) or gonadectomized when 90 days old (G). In the indirectly elicited muscle twitch, the LA from G rats was less sensitive to d-tubocurarine (0.1-1 microM) than control muscles (IC25: C = 0.30 microM, G = 0.46 microM). In G rats, the amplitude of neurally evoked endplate currents (EPC) was reduced by 70%, but the EPC time constant was not changed. Maximal junctional binding of [125I]alpha-bungarotoxin in the LA (C: 72.5 +/- 13.2 amol/endplate) was reduced by 1.8-fold in LA from G rats, with no change of the association rate constant (C: 5.64 +/- 1.29 10(6) M-1 min-1). The results indicate that testosterone deprivation reduces the junctional AChR density in the rat LA without modifying the binding properties of the receptor.

Efflux of acetylcholine in dimorphic skeletal muscle from castrated male rats.

The effect of testosterone on motor neurons of dimorphic muscles is demonstrable by the increased frequency of miniature end-plate potentials (mepp) and decreased end-plate acetylcholinesterase activity observed in castrated rats. No change occurs in induced acetylcholine (ACh) release. Although these muscles atrophy after castration there is no loss of muscle fibers. In the present study we reinvestigate the neuromuscular transmission in levator ani (LA) muscles from normal (N) adult (120 days) male rats and from rats castrated (C) 30 days before. The measurement of radioactive [3H]-choline was used to evaluate ACh release since it permits simultaneous estimation of quantal and non-quantal ACh release. The LA muscle was incubated with [3H]-choline (1 microCi/ml) for 30 min and ACh efflux was measured after washout. The basal release of [3H]-choline (dpm total tissue radioactivity-1 number of fibers-1) was 296 +/- 33 and 156 +/- 24 in N and C, respectively. Induced ACh release (25 Hz, 5 min) was the same in N and C (653.19 +/- 66.46 and 496.62 +/- 68.67, respectively). These results indicate that castration increased mepp frequency but reduced the total spontaneous release of ACh.

Efflux of acetylcholine in dimorphic skeletal muscle from castrated male rats

The effect of testosterone on motor neurons of dimorphic muscles is demostrable by increased frequency of miniature end-plate potentials (mepp) and decreased end-plate acetylcholinesterase activity observed in castrated rats. No change occurs in induced acetylcholine (ACh release. Although these muscles atrophy after castration there is no loss of muscle fibers. In the present study we reinvestigate the neuromuscular transmission in levator ani (LA) muscles from normal (N) adult (120 days) male rats and from rats castrated (C) 30 days before. The measurement of radioactive [3H]-choline was used to evaluate ACh release since it permits simultaneous estimation of quantal and non-quantal and non-quantal ACh release. The LA muscle was incubated with [3H]-choline (1µCi/ml) for 30 min and ACh efflux was measured after washout. The basal release of [3H]-choline (dpm total tissue radioactivity-1 number of fibers-1) was 296 ñ 33 and 156 ñ 24 in N and C, respectively. Induced ACh release (25 Hz, 5 min) was the same in N and C (653.19 ñ 66.46) and 496.62 ñ 68.67, respectively). These results indicate that castration increased nepp frequency buth reduced the total spontaneous release of ACh

Androgen regulation of the nicotinic acetylcholine receptor-ionic channel in a hormone-dependent skeletal muscle

The trophic influence of testosterone on the nicotinic acetylcholine receptor-ionic channel (AChR) was studied in the levator ani (LA) muscle of adult malr rats (120 days) intact (C) or gonadectomized when 90 days old (G). In the indirectly elicited muscle twitch, the LA from G rats was less sensitive to d-tubocurarine (0.1-1µM) than control muscles (IC25:C = 0.30µM,G=0.46µM). In G rats, the amplitude of neurally evoked endplate currents (EPC) was reduced by 70%, but the EPC time constant was not changed. Maximal junctional binding of [125I] alfa-bungarotoxin in the LA(C: 72.5 ñ 13.2 amol/endplate) was reduced by 18.8-fold in LA from G rats, with no change of the association rate constant (C: 5.64 ñ 1.29 10**6 M-1 min**-1). The results indicate that testosterone deprivation reduces the junctional AChR density in the rat LA without modifying the binding properties of the receptor

Castration atrophy and pharmacological reactivity of the rat coagulating gland.

1. The reactivity and sensitivity of the rat coagulating gland to acetylcholine, adrenaline, serotonin and barium chloride was studied 7, 15 and 30 days after castration. 2. The wet and the dry weight of the coagulating gland progressively decreased with time after castration. 3. Spontaneous contractions were observed in the coagulating gland 15 and 30 days after castration. 4. The maximum force developed per gram tissue was significantly higher than control for serotonin and barium chloride on the 7th day, while for acetylcholine and adrenaline the increase was observed only on the 15th day after castration. 5. The pD2 values for adrenaline, barium chloride and serotonin increased significantly 15 days after castration, whereas the change in sensitivity to acetylcholine was detected only 30 days after castration. 6. These results suggest that changes in reactivity and sensitivity of the rat coagulating gland caused by castration are not related only to muscle atrophy, since the time course of the development of the effects is different for the four agonists studied.