Foraging behaviour of a continental shelf marine predator, the grey seal (Halichoerus grypus), is associated with in situ, subsurface oceanographic conditions.

BACKGROUND: The heterogeneous oceanographic conditions of continental shelf ecosystems result in a three-dimensionally patchy distribution of prey available to upper-trophic level predators. The association of bio-physical conditions with movement patterns of large marine predators has been demonstrated in diverse taxa. However, obtaining subsurface data that are spatio-temporally relevant to the decisions made by benthically-foraging species can be challenging. METHODS: Between 2009 and 2015, grey seals were captured on Sable Island, Nova Scotia, Canada during summer and fall and instrumented with high-resolution archival GPS tags. These tags recorded location data as well as depth (m), temperature (°C), and light level measurements during dives, until animals returned to the haulout site to breed. Hidden Markov models were used to predict apparent foraging along movement tracks for 79 individuals (59 females, 20 males) every 3 h. In situ measurements were used to estimate chlorophyll-a concentration (mg m- 3) and temperature within the upper-water column (50 m) and temperature and depth at the bottom of dives. As chlorophyll-a could only be estimated from 10:00 to 14:00 AST for dive depths ≥50 m, we formulated two generalized linear mixed-effects models to test the association of predicted grey seal behavioural states with oceanographic conditions and phytoplankton biomass: the first representing conditions of the upper-water column likely to influence primary productivity, and a second model including environmental conditions encountered by grey seals at the bottom of dives, when seals were more likely to be foraging. RESULTS: Predicted grey seal behavioural states were associated with fine-scale chlorophyll-a concentrations and other environmental conditions they encountered across the continental shelf. In the Water Column Model, season had no influence on the probability of observing apparent foraging, but chlorophyll-a, upper-water column temperature, and sex did, with females having a greater probability of foraging than males. In the Bottom Conditions Model, again season had no influence on the probability of apparent foraging, but females were over twice as likely as males to be foraging. CONCLUSIONS: The results of this study highlight the value of in situ measurements of oceanographic properties that can be collected at high temporal resolution by animal-borne data loggers. These data provide insight into how inferred behavioural decisions made by large marine predators, such as the grey seal, may be influenced by fine-scale oceanographic conditions.

Sex-differences in fine-scale home-range use in an upper-trophic level marine predator.

BACKGROUND: The distribution of prey in the ocean is spatially and temporally patchy. How predators respond to this prey patchiness may have consequences on their foraging success, and thus physical condition. The recent ability to record fine-scale movements of marine animals combined with novel home-range analyses that incorporate the dimension of time should permit a better understanding of how individuals utilise different regions of space and the consequences on their foraging success. METHODS: Over a six-year study, we used T-LoCoH (Time-Local Convex Hull) home-range software to model archival GPS (Global Positioning System) data from 81 grey seals to investigate the fine-scale spatio-temporal use of space and the distribution of apparent foraging effort. Regions of home-ranges were classified according to the frequency of return visits (site fidelity) and duration of visits (intensity of use). Generalized linear mixed -effects models were used to test hypotheses on seasonal changes in foraging distribution and behaviour and the role of space-use and state on determining foraging success. RESULTS: Male grey seals had larger home-ranges and core areas than females, and both sexes showed a contraction in home-range and core area in fall leading up to the breeding season compared with summer. Heavier individuals had smaller core areas than lighter ones, suggesting access to higher quality habitat might be limited to those individuals with greater foraging experience and competitive ability. The size of the home-range or core area was not an important predictor of the rate of mass gain. A fine-scale spatio-temporal analysis of habitat use within the home-range provided evidence of intra-annual site fidelity at presumed foraging locations, suggesting predictably in prey distribution. Neither sex nor season were useful predictors for classifying behaviour. Rather, individual identity explained much of the variation in fine-scale behaviour. CONCLUSIONS: Understanding how upper-trophic level marine predators use space provides opportunities to explore the consequences of variation in foraging tactics and their success on fitness. Having knowledge of the drivers that shape this intraspecific variation can contribute toward predicting how these predators may respond to both natural and man-made environmental forcing.

Repeatability in lactation performance and the consequences for maternal reproductive success in gray seals.

In mammals, the most significant maternal effect on offspring growth during lactation is the ability of females to efficiently transfer milk energy to their neonates. However, despite the importance of the transfer of milk energy to both maternal and offspring fitness, nothing is known about the extent to which variation among females may be attributed to differences in individual quality or environmental variation in natural populations. We measured repeatability over multiple lactation periods in components of lactation performance in free-ranging, multiparous gray seal (Halichoerus grypus) females to examine to what extent variation among females in pup weaning mass may be attributed to inherent differences in their physiological capacity to deliver milk energy. Levels of repeatability were high for milk composition (r = 0.38-0.50), daily milk output (r = 0.46), and the duration of lactation (r = 0.57), demonstrating that there are consistent differences among females in these characters across lactations and that the overall capacity of gray seal females to deliver milk energy to their pups is characteristic of individuals. The repeatability in pup weaning mass (r = 0.48) was consistent with the values for the components of total milk energy output and suggests that, over a large proportion of their reproductive life, individual gray seal females will consistently wean pups with greater or lesser probabilities of survival. Our results suggest that inherent differences among females in their physiological capacity to deliver milk energy may be an important component of variation in individual quality and, thus, lifetime reproductive success in mammals. High levels of repeatability also suggest that components of milk energy transfer may have a significant heritable genetic basis.

Reproductive performance in grey seals: age-related improvement and senescence in a capital breeder.

1. Three hypotheses have been advanced to account for age-related improvement in performance: the selection hypothesis predicts improved due to the loss of lower quality phenotypes, the constraint hypothesis predicts individuals improve function, and the restraint hypothesis predicts younger individuals forego or reduce effort because of mortality risks. A decline in age-related performance (i.e. senescence) is predicted by mutation accumulation, antagonistic pleiotropy and disposable soma (wear and tear) hypotheses. 2. Using five measures of performance - birth rate, maternal and pup birth mass, pup weaning mass, weaning success and lactation length - we tested these hypotheses concerning age-related change in reproduction in 279 female grey seals (Halichoerus grypus), ages 4-42 years, over a 23-year period between 1983 and 2005 on Sable Island, Nova Scotia. These females produced 2071 pups. 3. Although body mass of primiparous females increased with age (4-7 years) birth mass of their pups did not, but pup weaning mass did. Second- and third-parity females of the same age as primiparous females gave birth to and weaned heavier pups. However, parity and age were dropped from models when maternal body mass was included. 4. The proportion of females giving birth varied significantly with maternal age, increasing in young females and then declining late in life. Weaning success rate also increased rapidly to about 8 years and subsequently declined in females > 32 years. 5. Generalized additive models indicated nonlinear changes in 3 day body mass (i.e. approximately birth mass) and weaning mass of pups as a function of maternal age, after accounting statistically for the effects of maternal body mass. Mixed-effects, repeated-measures models fitted to longitudinal data further supported the conclusion that pup birth mass and weaning mass vary nonlinearly with maternal age and indicated nonlinear changes in lactation duration. 6. We found some support for the constraint hypothesis, but our findings were not consistent with the selection hypothesis or the restraint hypothesis as the basis for improvement in reproductive performance. 7. Senescence was evident in multiple female and offspring traits, indicating the degeneration in function of several physiological systems as predicted by the disposable soma hypothesis.

Sexual segregation of seasonal foraging habitats in a non-migratory marine mammal.

Many animal species segregate by sex. Such segregation may be social in nature, or ecological, or both. Grey seals (Halichoerus grypus), like many large mammals, are sexually size dimorphic. In size dimorphic species, allometric differences in morphology, metabolic rate and reproductive costs are likely. Such differences may require the sexes to use different foraging strategies or different habitats. To investigate sexual segregation of habitat in grey seals, we used satellite tracks from 95 (male 46; female 49) adults breeding at Sable Island, Nova Scotia (44 degrees N, 60 degrees W) collected from 1995 to 2005. Location estimates were made from satellite fixes using a state-space movement model to estimate true locations and regularize them in time. Location estimates were used to calculate home range kernels of male and female habitat use each month. Month by sex kernel home ranges revealed striking differences and dynamics in habitat use between males and females on spatial scales broader than most terrestrial examples and at temporal and spatial resolutions rarely available for marine species. Differences were most pronounced just before (October-December) and immediately after breeding (February-March). During both periods, males primarily used areas along the continental shelf break, while females mainly used mid-shelf regions. Coupled with previously identified sex-specific seasonal patterns of energy storage, diving and diet, our findings suggest that males and females differ profoundly in their spatial foraging strategies. These differences may serve to maximize fitness by reducing intersexual competition during key foraging periods.

Stomach temperature telemetry reveals temporal patterns of foraging success in a free-ranging marine mammal.

1. We studied feeding frequency in free-ranging grey seals using stomach temperature telemetry to test if previously reported sex differences in the diving, movement and diet were reflected in the temporal pattern of foraging success. 2. Data were retrieved from 21 of 32 grey seals from 1999 to 2001, totalling 343 days and 555 feeding events, with individual record length varying from 2 to 40 days (mean: 16.33 +/- 2.67 days/seal). 3. Seals fed on 57.8 +/- 6.46% of days sampled and had an average of 1.7 +/- 0.26 meals per day, but individual variability was apparent in the temporal distribution of feeding as evidenced by high coefficients of variation (coefficient of variation = 69.0%). 4. Bout analysis of non-feeding intervals of six grey seals suggests that feeding intervals of individuals were varied and probably reflect differences in prey availability. Grey seals tended to have many single feeding events with long periods separating each event, as would be expected for a large carnivore with a batch-reactor digestive system. 5. We found significant sex differences in the temporal distribution of feeding. The number of feeding events per day was greater in males (2.2 +/- 0.4 vs. 1.0 +/- 0.2), as was time associated with feeding per day (56.6 +/- 5.8 min vs. 43.9 +/- 9.4 min). 6. The number of feeding events varied with time of day with the least number occurring during dawn. Feeding event size differed significantly by time of day, with greater meal sizes during the dawn and the smallest meals during the night. 7. The length of time between meals increased with the size of the previous meal, and was significantly less in males (541.4 +/- 63.5 min) than in females (1092.6 +/- 169.9 min). 8. These results provide new insight into the basis of sex differences in diving and diet in this large size-dimorphic marine predator.

The rate of fertilization in male mating tactics of the polygynous grey seal.

Studies using molecular markers have shown that some grey seal males may be gaining success through exhibiting alternative mating tactics. We estimated the probability of fertilization success of grey seal males exhibiting the primary tactic of female defence and one alternative tactic of mating with departing females on Sable Island, Nova Scotia, Canada, during the breeding seasons of 1997-2002. Although the fertilization rate of the primary tactic (27-43%) was greater than that of the alternative tactic (10-12%), these low rates indicate the potential fitness value of alternative mating tactics in this size-dimorphic pinniped species.

Novel ceramide analogues display selective cytotoxicity in drug-resistant breast tumor cell lines compared to normal breast epithelial cells.

The sphingolipid ceramide is involved in diverse cell signaling pathways related to proliferation and differentiation. Elevated ceramide also triggers apoptosis. Synthetic ceramide derivatives have been shown to be cytotoxic to tumors, yet few studies have evaluated whether cytotoxicity of synthetic ceramides is selective for tumor cells. We have evaluated the cytotoxic potency of several novel ceramide analogues in the drug-resistant breast tumor cell lines, SKBr3 and MCF-7/Adr, and compared their cytotoxicity in normal breast epithelial cells. Cytotoxicity was assessed using release of lactate dehydrogenase into the culture medium. (2S, 3S)-3-(6'-Dodecylpyridin-2'-yl)-2-butanoylamidopropane-1,3-diol (pyridine-C4-ceramide) produced non-selective cytotoxicity across the three cell types (EC50= 12.8-16.7 microM, at 24 hr). However, 2S,5R-2-(octanoylamido-(3E))-octadecene-1,5-diol (5R-OH-3E-C8-ceramide), (2S,3R)-2-(N-adamantoyl)-(4E)-octadecen-1,3-diol (adamantyl-ceramide), and (2S,3R)-3-(3'-dodecylphenyl)-2-butanoylamidopropane-1,3-diol (benzene-C4-ceramide) exhibited increased cytotoxicity in the tumor cell lines compared to the normal breast epithelial cells. The EC50 values (microM) at 24 hr for these compounds in SKBr3 cells, MCF-7/Adr cells, and normal breast epithelial cells, respectively, were as follows: 5R-OH-3E-C8-ceramide, 18.3, 21.2 and 58.7; adamantyl-ceramide, 10.9, 24.9 and >100; benzene-C4-ceramide, 18.9, 45.5 and >100. At a concentration of 30 microM, the fold increase in cytotoxicity in breast tumor cell lines compared with normal breast epithelial cells was as follows: 5R-OH-3E-C8-ceramide, 23.7 and 19; adamantyl-ceramide, 11.2 and 10.3 and benzene-C4-ceramide, 79.3 and 77.2, for SKBr3 and MCF-7/Adr cells, respectively. Possible mechanisms accounting for selectivity are discussed. Ceramide analogues with relatively selective toxicity against tumor cells may have potential as therapeutic agents. Elucidating the mechanisms of selective cytotoxicity could identify novel targets that may lead to development of anti-neoplastic agents with a higher therapeutic index.

Weaning mass affects changes in body composition and food intake in harbour seal pups during the first month of independence.

In phocid seals, the transition to nutritional independence is abrupt, with females abandoning their offspring after weaning and returning to sea. We hypothesized that body size at weaning may play an important role in the nature of this transition. We studied the changes in body composition and water flux of newly weaning harbour seals over the first 4-6 wk postweaning. Thirty-three pups were dosed with deuterium oxide to estimate total body water (TBW) and a subset of 24 was dosed twice to estimate changes in body composition and water flux. All pups lost body mass over the study period, but TBW increased during the period of mass loss, indicating continued lean tissue growth. Combined data from this and our early study indicated that heavy (>median mass) pups were relatively fatter (41.0% vs. 37.1%) and had significantly greater total body energy at weaning than did light (< or = median mass) pups. Percentage TBW declined linearly over time in light pups but was constant in heavy pups for the first 19 d postweaning and then declined linearly. Both the temporal pattern and composition of mass loss differed between light and heavy pups. Estimated food intake increased in the second 2 wk of study compared to the first 2 wk, in both light and heavy pups, reflecting increased foraging success but at levels still insufficient to meet daily expenditures of most individuals.

Synthesis and biological evaluation of tropane-like 1-[2-[bis(4-fluorophenyl)methoxy]ethyl]-4-(3-phenylpropyl)piperazine (GBR 12909) analogues.

We have prepared azabicyclo[3.2.1] derivatives (C-3-substituted tropanes) that bind with high affinity to the dopamine transporter and inhibit dopamine reuptake. Within the series, 3-[2-[bis-(4-fluorophenyl)methoxy]ethylidene]-8-methyl-8-azabicyclo[3.2.1]octane (8) was found to have the highest affinity and selectivity for the dopamine transporter. These azabicyclo[3.2.1] (bridged piperidine) series of compounds differ from the well-known benztropines by a 2-carbon spacer between C-3 and a diarylmethoxy moiety. Interestingly, these new compounds demonstrated a much lower affinity for the muscarinic-1 site, at least a 100-fold decrease compared to benztropine. Replacing N-methyl with N-phenylpropyl in two of the compounds resulted in a 3-10-fold increase in binding affinity for the dopamine transporter. However, those compounds lost selectivity for the dopamine transporter over the serotonin transporter. Replacement of the ether oxygen in the diarylmethoxy moiety with a nitrogen atom gave relatively inactive amines, indicating the important role which is played by the ether oxygen in transporter binding. Reduction of the C-3 double bond in 8 gave 3 alpha-substituted tropanes, as shown by X-ray crystallographic analyses of 11, 12, and 19. The 3 alpha-substituted tropanes had lower affinity and less selectivity than the comparable unsaturated ligands.

Rimcazole analogs attenuate the convulsive effects of cocaine: correlation with binding to sigma receptors rather than dopamine transporters.

Cocaine interacts with dopamine transporters and sigma receptors at concentrations that are achievable in vivo, suggesting that they may both be viable targets for the development of anti-cocaine agents. Rimcazole binds to both of these targets and also attenuates cocaine-induced locomotor activity and sensitization. To further characterize the mechanism(s) underlying the attenuation of cocaine-induced convulsions and lethality, rimcazole and three analogs (SH3/24, SH2/21, SH1/57), with a range of affinities for dopamine transporters and sigma receptors, were evaluated. The highly selective and potent sigma receptor ligand LR176 was used as a reference. Competition binding studies confirmed that the rank order of the compounds at dopamine transporters vs. sigma receptors differed, thus enabling a correlation between the relative anti-cocaine activities of the compounds in behavioral studies and their affinities for dopamine transporters vs. sigma receptors. In behavioral studies, male Swiss Webster mice were pre-treated with one of the compounds (0-60 mg/kg, i.p.), then challenged 15 min later with either a convulsive (60 mg/kg, i.p.) or lethal (125 mg/kg, i.p.) dose of cocaine. When the compounds were ranked according to their protective effect, there was a significant correlation between their anticonvulsant actions and their affinities for sigma receptors, but not dopamine transporters. Although the rimcazole analogs were ineffective against the lethal effects of cocaine, the selective sigma receptor ligand LR176 provided significant protection. These data thus suggest that sigma receptors may mediate some of the toxic effects associated with cocaine and that sigma receptor antagonists may be developed as pharmacotherapeutic agents for this application.

Conformationally restricted analogs of BD1008 and an antisense oligodeoxynucleotide targeting sigma1 receptors produce anti-cocaine effects in mice.

Cocaine's ability to interact with sigma receptors suggests that these proteins mediate some of its behavioral effects. Therefore, three novel sigma receptor ligands with antagonist activity were evaluated in Swiss Webster mice: BD1018 (3S-1-[2-(3,4-dichlorophenyl)ethyl]-1,4-diazabicyclo[4.3.0]nonane), BD1063 (1-[2-(3,4-dichlorophenyl)ethyl]-4-methylpiperazine), and LR132 (1R,2S-(+)-cis-N-[2-(3,4-dichlorophenyl)ethyl]-2-(1-pyrrolidinyl)cyclohexylamine). Competition binding assays demonstrated that all three compounds have high affinities for sigma1 receptors. The three compounds vary in their affinities for sigma2 receptors and exhibit negligible affinities for dopamine, opioid, GABA(A) and NMDA receptors. In behavioral studies, pre-treatment of mice with BD1018, BD1063, or LR132 significantly attenuated cocaine-induced convulsions and lethality. Moreover, post-treatment with LR132 prevented cocaine-induced lethality in a significant proportion of animals. In contrast to the protection provided by the putative antagonists, the well-characterized sigma receptor agonist di-o-tolylguanidine (DTG) and the novel sigma receptor agonist BD1031 (3R-1-[2-(3,4-dichlorophenyl)ethyl]-1,4-diazabicyclo[4.3.0]nonane) each worsened the behavioral toxicity of cocaine. At doses where alone, they produced no significant effects on locomotion, BD1018, BD1063 and LR132 significantly attenuated the locomotor stimulatory effects of cocaine. To further validate the hypothesis that the anti-cocaine effects of the novel ligands involved antagonism of sigma receptors, an antisense oligodeoxynucleotide against sigma1 receptors was also shown to significantly attenuate the convulsive and locomotor stimulatory effects of cocaine. Together, the data suggests that functional antagonism of sigma receptors is capable of attenuating a number of cocaine-induced behaviors.

N-alkyl substituted analogs of the sigma receptor ligand BD1008 and traditional sigma receptor ligands affect cocaine-induced convulsions and lethality in mice.

Cocaine binds to sigma receptors with comparable affinity to its well-established interaction with dopamine transporters. Previous studies have shown BD1008 (N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1-pyrrolidinyl)ethylamine) to have high affinity and selectivity for sigma receptors, and to additionally attenuate the locomotor stimulatory effects of cocaine. Therefore, in the present study, three N-alkyl substituted analogs of BD1008 were characterized in receptor binding and behavioral studies: BD1060 (N-[2-(3,4-dichlorophenyl)ethyl]-2-(1-pyrrolidinyl)ethylamine), BD1067 (N-[2-(3,4-dichlorophenyl)ethyl]-N-ethyl-2-(1-pyrrolidinyl)ethylamine), and BD1052 (N-[2-(3,4-dichlorophenyl)ethyl]-N-allyl-2-(1-pyrrolidinyl)ethylamine). Similarly to BD1008, all three analogs exhibited high affinity and selectivity for sigma receptors. In behavioral studies, BD1008, BD1060 or BD1067 attenuated cocaine-induced convulsions and lethality in Swiss Webster mice. The protective effects appear to be mediated through sigma receptor antagonism because traditional sigma receptor antagonists with high to moderate affinity for these receptors also attenuated the behavioral toxicity of cocaine. In contrast, traditional and novel sigma receptor agonists such as di-o-tolylguanidine and BD1052 worsened the behavioral toxicity of cocaine. To further characterize the actions of the N-alkyl substituted compounds, they were microinjected into the rat red nucleus, a functional assay of sigma receptor activity, where they produced agonist vs. antagonist actions that were consistent with their effects on cocaine-induced behaviors. Together, the data demonstrate that BD1008, BD1060 or BD1067 can attenuate the behavioral toxicity of cocaine, most likely through functional antagonism of sigma receptors.

Metabolic compensation during high energy output in fasting, lactating grey seals (Halichoerus grypus): metabolic ceilings revisited.

Lactation is the most energetically expensive period for female mammals and is associated with some of the highest sustained metabolic rates (SusMR) in vertebrates (reported as total energy throughput). Females typically deal with this energy demand by increasing food intake and the structure of the alimentary tract may act as the central constraint to ceilings on SusMR at about seven times resting or standard metabolic rate (SMR). However, demands of lactation may also be met by using a form of metabolic compensation such as reducing locomotor activities or entering torpor. In some phocid seals, cetaceans and bears, females fast throughout lactation and thus cannot offset the high energetic costs of lactation through increased food intake. We demonstrate that fasting grey seal females sustain, for several weeks, one of the highest total daily energy expenditures (DEE; 7.4 x SMR) reported in mammals, while progressively reducing maintenance metabolic expenditures during lactation through means not explained by reduction in lean body mass or behavioural changes. Simultaneously, the energy-exported in milk is progressively increased, associated with increased lipoprotein lipase activity in the mammary gland, resulting in greater offspring growth. Our results suggest that females use compensatory mechanisms to help meet the extraordinary energetic costs of lactation. Additionally, although the concepts of SusMR and ceilings on total DEE may be somewhat different in fasting lactating species, our data on phocid seals demonstrate that metabolic ceilings on milk energy output, in general, are not constrained by the same kind of peripheral limitations as are other energy-consuming tissues. In phocid seals, the high ceilings on DEE during lactation, coupled with metabolic compensation, are undoubtedly important factors enabling shortened lactation.

Seasonal changes in buoyancy and diving behaviour of adult grey seals.

Phocid seals go through dramatic seasonal changes in body mass and composition as a result of the spatial and temporal separation of foraging, reproduction and moulting. These changes in body fat content and body mass result in seasonal changes in buoyancy, which in turn may influence diving behaviour. We examined the longitudinal changes in buoyancy and diving behaviour of 14 adult grey seals (Halichoerus grypus) during two periods that represent maximal contrast in body mass and composition. During both the post-moulting (PM) and pre-breeding (PB) periods, grey seals were negatively buoyant. However, buoyancy increased by 47.9 % between the PM and PB periods. Descent rate was significantly faster during the PM period (1.0+/-0.07 m s(-1)) than during the PB period (0.7+/-0.06 m s(-1)), suggesting that seals were aided by negative buoyancy during the downward portion of dives. Ascent rate was also significantly faster during the PM period (0.8+/-0.06 m s(-1)) than during the PB period (0.6+/-0.05 m s(-1)), contradicting the prediction that more buoyant animals should ascend faster. The effects of drag could not account for this discrepancy. Dive depth and surface interval between dives did not differ significantly between the two periods. Similarly, the distribution of dive shapes used by individuals did not differ between the two periods. However, dive duration was significantly longer during the PB period than during the PM period (5.5+/-0.25 min compared with 4.4+/-0.24 min, respectively) as was time spent at the bottom of the dive (3.1+/-0.22 min compared with 2.5+/-0.15 min, respectively).

Sigma receptors: recent advances and new clinical potentials.

Several recent advances are leading to a better understanding of sigma receptors. Here we focus on our recent findings regarding cellular functions of sigma-2 receptors and discuss their possible clinical implications. Agonists at sigma-2 receptors induced changes in cell morphology and apoptosis in various cell types. Sigma-2 receptor activation produced both transient and sustained increases in [Ca++]i, derived from different intracellular stores. These changes in [Ca++]i and cytotoxic effects are mediated by intracellular sigma-2 receptors. Sigma-2 agonists induced apoptosis in drug-resistant cancer cells, enhanced the potency of DNA damaging agents, and down-regulated expression of p-glycoprotein mRNA. Thus, sigma-2 receptor agonists may be useful in treatment of drug-resistant cancers. Sigma radioligands have been used in tumor imaging. We also discuss how sigma-2 antagonists might prevent the irreversible motor side effects of typical neuroleptics. Sigma-2 receptors may subserve a novel signalling pathway to apoptosis, involved in regulation of cell proliferation and/or viability.

Effect of a low-Fat diet on body composition and blubber fatty acids of captive juvenile harp seals (Phoca groenlandica).

We investigated the effects of a change from a high-fat diet to a low-fat diet of differing fatty acid (FA) composition on the body composition and blubber FA of five captive juvenile harp seals. Seals that had been maintained for 1 yr on a diet of Atlantic herring (>/=9% fat) were switched to a diet of Atlantic pollock (1. 7% fat) for 30 d. On days 0, 14, and 30, mass and body composition (using isotope dilution) were measured, and blubber biopsies (5 cmx6 mm) were taken for FA analysis. Fat accounted for 38%-49% of body mass at the start of the experiment. When switched to the pollock diet, and despite food intakes averaging 6.5 kg/d (32.3 MJ/d), body fat declined by an average of 6.4 kg or by 32% over the 30-d experiment. In contrast, body protein increased in direct relation to protein intake (r2=0.836, P=0.030). Despite substantial loss of body fat, blubber FA signature changed significantly to reflect the changes in dietary intake of FA, and the deposition of FA was quantifiably predictable. Our results suggest that young growing phocids are unable to maintain body fat stores on low-fat diets even when protein intakes are high. This may have significant implications for juvenile pinniped survival in the wild. In addition, turnover and deposition of dietary FA in blubber takes place in nonfattening seals.

Modulation of cellular calcium by sigma-2 receptors: release from intracellular stores in human SK-N-SH neuroblastoma cells.

Human SK-N-SH neuroblastoma cells expressed sigma-1 and sigma-2 receptors with similar pharmacological profiles to those of rodent-derived tissues, although sigma-2 receptors exhibited some affinity differences that might suggest heterogeneity or species differences. Structurally diverse sigma ligands produced two types of increases in intracellular (cytosolic) Ca(2+) concentration ([Ca(2+)](i)) in these cells. CB-64D, CB-64L, JL-II-147, BD737, LR172, BD1008, haloperidol, reduced haloperidol, and ibogaine all produced an immediate, dose-dependent, and transient rise in [Ca(2+)](i). Sigma-inactive compounds structurally similar to the most active sigma ligands and ligands for several neurotransmitter receptors produced little or no effect. The high activity of CB-64D and ibogaine (sigma-2-selective ligands) compared with the low activity of (+)-pentazocine and other (+)-benzomorphans (sigma-1-selective ligands), in addition to enantioselectivity for CB-64D over CB-64L, strongly indicated mediation by sigma-2 receptors. The effect of CB-64D and BD737 was blocked by the sigma antagonists BD1047 and BD1063, further confirming specificity as a receptor-mediated event. The transient rise in [Ca(2+)](i) occurred in the absence of extracellular Ca(2+) and was completely eliminated by pretreatment of cells with thapsigargin. Thus, sigma-2 receptors stimulate a transient release of Ca(2+) from the endoplasmic reticulum. Prolonged exposure of cells to sigma-receptor ligands resulted in a latent and sustained rise in [Ca(2+)](i), with a pharmacological profile identical to that of the transient rise. This sustained rise in [Ca(2+)](i) was affected by neither the removal of extracellular Ca(2+) nor thapsigargin pretreatment, suggesting latent sigma-2 receptor-induced release from thapsigargin-insensitive intracellular Ca(2+) stores. Sigma-2 receptors may use Ca(2+) signals in producing cellular effects.

A sigma-1 receptor selective analogue of BD1008. A potential substitute for (+)-opioids in sigma receptor binding assays.

A simple, achiral monoamine sigma-1 (sigma1) receptor selective ligand (sigma2Ki/sigma1Ki>2000) is described, which could replace the chiral (+)-pentazocine or dextrallorphan as a sigma1 masking agent in sigma2 binding assays.