Forward shift of feeding buzz components of dolphins and belugas during associative learning reveals a likely connection to reward expectation, pleasure and brain dopamine activation.

For many years, we heard sounds associated with reward from dolphins and belugas. We named these pulsed sounds victory squeals (VS), as they remind us of a child's squeal of delight. Here we put these sounds in context with natural and learned behavior. Like bats, echolocating cetaceans produce feeding buzzes as they approach and catch prey. Unlike bats, cetaceans continue their feeding buzzes after prey capture and the after portion is what we call the VS. Prior to training (or conditioning), the VS comes after the fish reward; with repeated trials it moves to before the reward. During training, we use a whistle or other sound to signal a correct response by the animal. This sound signal, named a secondary reinforcer (SR), leads to the primary reinforcer, fish. Trainers usually name their whistle or other SR a bridge, as it bridges the time gap between the correct response and reward delivery. During learning, the SR becomes associated with reward and the VS comes after the SR rather than after the fish. By following the SR, the VS confirms that the animal expects a reward. Results of early brain stimulation work suggest to us that SR stimulates brain dopamine release, which leads to the VS. Although there are no direct studies of dopamine release in cetaceans, we found that the timing of our VS is consistent with a response after dopamine release. We compared trained vocal responses to auditory stimuli with VS responses to SR sounds. Auditory stimuli that did not signal reward resulted in faster responses by a mean of 151 ms for dolphins and 250 ms for belugas. In laboratory animals, there is a 100 to 200 ms delay for dopamine release. VS delay in our animals is similar and consistent with vocalization after dopamine release. Our novel observation suggests that the dopamine reward system is active in cetacean brains.

A case series of patients with lamotrigine toxicity at one center from 2003 to 2012.

INTRODUCTION: Lamotrigine is a phenyltriazine compound that inhibits voltage-gated sodium channels, decreasing release of glutamate and aspartate, and inhibits serotonin, norepinephrine and dopamine reuptake. Reports of toxicity in the literature are limited to case reports and primarily involve coingestants. This case series is intended to report the clinical manifestations of lamotrigine toxicity. METHODS: This retrospective case series from 2003 to 2012 studies the effects of lamotrigine toxicity when not confounded by coingestants. Admission records at an inpatient toxicology center were reviewed for lamotrigine-only exposure based on history with supporting laboratory data when available. After identification, these charts were reviewed again to characterize vital signs, neurological examination findings, specific laboratory and electrocardiography parameters, and complications. RESULTS: Fifty-seven patients were identified with possible lamotrigine toxicity. Nine patients, including three toddlers, had lamotrigine-only ingestions. Three of these patients had seizures, four were hypertensive, five were tachycardic, and four experienced tachypnea. Mental status was altered in all nine (depressed (n = 4), agitated (n = 5) or both (n = 3)). Five patients were hyperreflexic and experienced intermittent myoclonus, and two had inducible clonus. On electrocardiogram, two patients experienced QRS prolongation (114-116 ms), and four had QTc prolongation (463-586 ms). No patient had life-threatening symptoms or signs. Serum levels of lamotrigine were available in seven patients, and averaged 35.4 mg/L (17-90 mg/L). The therapeutic range for sLTG is 3-14 mg/L. CONCLUSIONS: Lamotrigine toxicity manifested with minor-moderate neurologic and/or electrocardiographic effects. Toxicity reflects the known pharmacologic actions of lamotrigine: serotonin, norepinephrine and dopamine reuptake inhibition, and sodium channel blockade.

Reversible mechanical actuation of elastomeric nanopores.

Mechanical resizing of individual nanopores in a thermoplastic polyurethane elastomer has been characterized. Specimen nanopores were conical, with smaller hole dimensions of the order of tens to hundreds of nanometres. Electrophoretic current measurements show that the estimated nanopore radius can be reversibly actuated over an order of magnitude by stretching and relaxing the elastomer. Within a working range of stretching, current is proportional to specimen extension to the power of a constant, n, which ranges from 0.9 to 2.3 for different specimens. The data indicate that scaling of the effective pore radius is super-affine. At strains below the working range, the pore size is relatively unresponsive to stretching. Macroscopic elastomer extension has been related to local radial strain (50-250 µm from the pore) using optical microscopy. Scanning electron microscopy and atomic force microscopy have been used to observe membrane surface features.

The plastid DNA of the malaria parasite Plasmodium falciparum is replicated by two mechanisms.

In common with other apicomplexan parasites, Plasmodium falciparum, a causative organism of human malaria, harbours a residual plastid derived from an ancient secondary endosymbiotic acquisition of an alga. The function of the 35 kb plastid genome is unknown, but its evolutionary origin and genetic content make it a likely target for chemotherapy. Pulsed field gel electrophoresis and ionizing radiation have shown that essentially all the plastid DNA comprises covalently closed circular monomers, together with a tiny minority of linear 35 kb molecules. Using two-dimensional gels and electron microscopy, two replication mechanisms have been revealed. One, sensitive to the topoisomerase inhibitor ciprofloxacin, appears to initiate at twin D-loops located in a large inverted repeat carrying duplicated rRNA and tRNA genes, whereas the second, less drug sensitive, probably involves rolling circles that initiate outside the inverted repeat.

Assessment of dolphin (Tursiops truncatus) auditory sensitivity and hearing loss using jawphones.

Devices known as jawphones have previously been used to measure interaural time and intensity discrimination in dolphins. This study introduces their use for measuring hearing sensitivity in dolphins. Auditory thresholds were measured behaviorally against natural background noise for two bottlenose dolphins (Tursiops truncatus); a 14-year-old female and a 33-year-old male. Stimuli were delivered to each ear independently by placing jawphones directly over the pan bone of the dolphin's lower jaw, the assumed site of best reception. The shape of the female dolphin's auditory functions, including comparison measurements made in the free field, favorably matches that of the accepted standard audiogram for the species. Thresholds previously measured for the male dolphin at 26 years of age indicated a sensitivity difference between the ears of 2-3 dB between 4-10 kHz, which was considered unremarkable at the time. Thresholds for the male dolphin reported in this study suggest a high-frequency loss compared to the standard audiogram. Both of the male's ears have lost sensitivity to frequencies above 55 kHz and the right ear is 16-33 dB less sensitive than the left ear over the 10-40 kHz range, suggesting that males of the species may lose sensitivity as a function of age. The results of this study support the use of jawphones for the measurement of dolphin auditory sensitivity.

The in vivo conformation of the plastid DNA of Toxoplasma gondii: implications for replication.

The Phylum Apicomplexa comprises thousands of obligate intracellular parasites, some of which cause serious disease in man and other animals. Though not photosynthetic, some of them, including the malaria parasites (Plasmodium spp.) and the causative organism of Toxoplasmosis, Toxoplasma gondii, possess a remnant plastid partially determined by a highly derived residual genome encoded in 35 kb DNA. The genetic maps of the plastid genomes of these two organisms are extremely similar in nucleotide sequence, gene function and gene order. However, a study using pulsed field gel electrophoresis and electron microscopy has shown that in contrast to the malarial version, only a minority of the plastid DNA of Toxoplasma occurs as circular 35 kb molecules. The majority consists of a precise oligomeric series of linear tandem arrays of the genome, each oligomer terminating at the same site in the genetic map, i.e. in the centre of a large inverted repeat (IR) which encodes duplicated tRNA and rRNA genes. This overall topology strongly suggests that replication occurs by a rolling circle mechanism initiating at the centre of the IR, which is also the site at which the linear tails of the rolling circles are processed to yield the oligomers. A model is proposed which accounts for the quantitative structure of the molecular population. It is relevant that a somewhat similar structure has been reported for at least three land plant chloroplast genomes.

An alteration in concatameric structure is associated with efficient segregation of plasmids in transfected Plasmodium falciparum parasites.

Transfection of the human malaria parasite Plasmodium falciparum is currently performed with circularised plasmids that are maintained episomally in parasites under drug selection but which are rapidly lost when selection pressure is removed. In this paper, we show that in instances where gene targeting is not favoured, transfected plasmids can change to stably replicating forms (SRFs) that are maintained episomally in the absence of drug selection. SRF DNA is a large concatamer of the parental plasmid comprising at least nine plasmids arranged in a head-to-tail array. We show as well that the original unstable replicating forms (URFs) are also present as head-to-tail concatamers, but only comprise three plasmids. Limited digestion and gamma irradiation experiments revealed that while URF concatamers are primarily circular, as expected, SRF concatamers form a more complex structure that includes extensive single-stranded DNA. No evidence of sequence rearrangement or additional sequence was detected in SRF DNA, including in transient replication experiments designed to select for more efficiently replicating plasmids. Surprisingly, these experiments revealed that the bacterial plasmid alone can replicate in parasites. Together, these results imply that transfected plasmids are required to form head-to-tail concatamers to be maintained in parasites and implicate both rolling-circle and recombination-dependent mechanisms in their replication.

The diving physiology of bottlenose dolphins (Tursiops truncatus). II. Biomechanics and changes in buoyancy at depth.

During diving, marine mammals must balance the conservation of limited oxygen reserves with the metabolic costs of swimming exercise. As a result, energetically efficient modes of locomotion provide an advantage during periods of submergence and will presumably increase in importance as the animals perform progressively longer dives. To determine the effect of a limited oxygen supply on locomotor performance, we compared the kinematics and behavior of swimming and diving bottlenose dolphins. Adult bottlenose dolphins (Tursiops truncatus) were trained to swim horizontally near the water surface or submerged at 5 m and to dive to depths ranging from 12 to 112 m. Swimming kinematics (preferred swimming mode, stroke frequency and duration of glides) were monitored using submersible video cameras (Sony Hi-8) held by SCUBA divers or attached to a pack on the dorsal fin of the animal. Drag and buoyant forces were calculated from patterns of deceleration for horizontally swimming and vertically diving animals. The results showed that dolphins used a variety of swimming gaits that correlated with acceleration. The percentage of time spent gliding during the descent phase of dives increased with depth. Glide distances ranged from 7.1+/-1.9 m for 16 m dives to 43.6+/-7.0 m (means +/- s.e.m.) for 100 m dives. These gliding patterns were attributed to changes in buoyancy associated with lung compression at depth. By incorporating prolonged glide periods, the bottlenose dolphin realized a theoretical 10-21 % energetic savings in the cost of a 100 m dive in comparison with dives based on neutral buoyancy models. Thus, modifying locomotor patterns to account for physical changes with depth appears to be one mechanism that enables diving mammals with limited oxygen stores to extend the duration of a dive.

Classification of dolphin echolocation clicks by energy and frequency distributions.

Dolphins demonstrate an adaptive control over echolocation click production, but little is known of the manner or degree with which control is exercised. Echolocation clicks (N approximately 30,000) were collected from an Atlantic bottlenose dolphin (Tursiops truncatus) performing object discrimination tasks in order to investigate differential click production. Seven categories of clicks were identified using the spectral conformation and relative position of -3 and -10 dB peaks. A counterpropagation network utilizing 16 inputs, 50 hidden units, and 8 output units was trained to classify clicks using the same spectral variables. The network classified novel clicks with 92% success. Additional echolocation clicks (N > 24,000) from two other dolphins were submitted to the network for classification. Classified echolocation clicks were analyzed for animal specific differences, changes in predominant click type within click trains, and task-related specificity. Differences in animal and task performance may influence click type and click train length.

Signal processing of the echo signatures returned by submerged shells insonified by dolphin "clicks:" active classification.

A large set of dolphin-emitted acoustic pulses ("echolocation clicks") have been examined, which were reflected from various elastic shells that were suspended, underwater, 4.5 m in front of the animal in a large test site in Kaneohe Bay, Hawaii. A carefully instrumented analog-to-digital system continuously captured the emitted clicks and also the returned, backscattered echoes (A/D conversion at 500 kHz). Using standard conditioning techniques and food reinforces, the dolphin is taught to push an underwater paddle when the "correct" target-the one he has been trained to identify-is presented to him. He communicates his consistently correct identifying choices in this manner. Many echoes returned by three types of cylindrical shells in both the time and frequency domains as well as in the joint time-frequency (t-f) domain, by means of Wigner-type distributions have been examined. It will be shown exactly how specific features observable in these displays are directly related to the physical characteristics of the shells. This processing takes advantage of certain fundamental resonance principles to show which echo features contain information about the size, shape, wall thickness, and material composition of both the shell and its filler substance. In the same fashion that these resonance features give the identifying characteristics of each shell, it is believed they may also give them to the dolphin. These echo features may allow him to extract the target properties by inspection without any need for computations. It is claimed that this may be the fundamental physical explanation of the dolphin's amazing target ID feats, upon which they base their recognition choices. This claim may be substantiated by the detailed analysis of many typical echoes returned by various shells, when they are interrogated by several dolphins. Thus far, this analysis of many echoes from many shells has only been carried out for a single dolphin.

Complete gene map of the plastid-like DNA of the malaria parasite Plasmodium falciparum.

Malaria parasites, and other parasitic protists of the Phylum Apicomplexa, carry a plastid-like genome with greatly reduced sequence complexity. This 35 kb DNA circle resembles the plastid DNA of non-photosynthetic plants, encoding almost exclusively components involved in gene expression. The complete gene map described here includes genes for duplicated large and small subunit rRNAs, 25 species of tRNA, three subunits of a eubacterial RNA polymerase, 17 ribosomal proteins, and a translation elongation factor. In addition, it codes for an unusual member of the Clp family of chaperones, as well as an open reading frame of unknown function found in red algal plastids. Transcription is polycistronic. This plastid-like DNA molecule is conserved in several genera of apicomplexans and is conjectured to have been acquired by an early progenitor of the Phylum by secondary endosymbiosis. The function of the organelle (plastid) carrying this DNA remains obscure, but appears to be specified by genes transferred to the nucleus.

Recombination associated with replication of malarial mitochondrial DNA.

Mitochondrial DNA of the malarial parasite Plasmodium falciparum comprises approximately 20 copies per cell of a 6 kb genome, arranged mainly as polydisperse linear concatemers. In synchronous blood cultures, initiation of mtDNA replication coincides with the start of the 4-5 doublings in nuclear DNA that mark the reproductive phase of the erythrocytic cycle. We show that mtDNA replication coincides with a recombination process reminiscent of the replication mechanism used by certain bacteriophages and plasmids. The few circular forms of mtDNA which are also present do not replicate by a theta mechanism, but are themselves the product of recombination, and we propose they undergo rolling circle activity to generate the linear concatemers.

Phylogenetic analysis of the rpoB gene from the plastid-like DNA of Plasmodium falciparum.

Malaria and other Apicomplexan parasites harbour two extrachromosomal DNAs. One is mitochondrial and the other is a 35-kb circle with some plastid-like features but whose provenance and function is unknown. In addition to genes for rRNAs, tRNAs and ribosomal proteins, the 35-kb circular DNA of Plasmodium falciparum carries an rpoBC operon which encodes subunits of a eubacteria-like RNA polymerase. The phylogenetic analysis of the complete rpoB sequence presented here supports our inference that the 35-kb circle is the remnant of a plastid genome.

Target detection, shape discrimination, and signal characteristics of an echolocating false killer whale (Pseudorca crassidens).

This study demonstrated the ability of a false killer whale (Pseudorca crassidens) to discriminate between two targets and investigated the parameters of the whale's emitted signals for changes related to test conditions. Target detection performance comparable to the bottlenose dolphin's (Tursiops truncatus) has previously been reported for echolocating false killer whales. No other echolocation capabilities have been reported. A false killer whale, naive to conditioned echolocation tasks, was initially trained to detect a cylinder in a "go/no-go" procedure over ranges of 3 to 8 m. The transition from a detection task to a discrimination task was readily achieved by introducing a spherical comparison target. Finally, the cylinder was successfully compared to spheres of two different sizes and target strengths. Multivariate analyses were used to evaluate the parameters of emitted signals. Duncan's multiple range tests showed significant decreases (df = 185, p less than 0.05) in both source level and bandwidth in the transition from detection to discrimination. Analysis of variance revealed a significant decrease in the number of clicks over test conditions [F(5.26) = 5.23, p less than 0.0001]. These data suggest that the whale relied on cues relevant to target shape as well as target strength, that changes in source level and bandwidth were task-related, that the decrease in clicks was associated with learning experience, and that Pseudorca's ability to discriminate shapes using echolocation may be comparable to that of Tursiops truncatus.

Detection of complex echoes in noise by an echolocating dolphin.

Dolphins echolocate with short broadband acoustic signals that have good time resolution properties. Received echoes are often complex, with many resolvable highlights or components caused by reflection of the incident signal from external and internal boundaries of a target and from different propagational modes within a target. A series of experiments was performed to investigate how dolphins perceive complex echoes. Echoes were produced by a microprocessor-controlled electronic target simulator that captured each emitted click and retransmitted the signal back to the animal after an appropriate time delay. The use of this "phantom" target allowed for precise control of the number of highlights, the time separation between highlights, and the relative amplitudes of highlights in the simulated echoes. An echolocating dolphin was trained to perform a target detection task in the presence of masking noise using these phantom echoes. The properties of simulated echoes were systematically varied, and corresponding shifts in the dolphin's detection threshold were observed, allowing for inferences of how the dolphin perceived echoes. The dolphin performed like an energy detector with an integration time of approximately 264 microseconds.

Zirconium molybdate gel as a generator for technetium-99m--I. The concept and its evaluation.

A new 99mTc generator for the preparation of radiopharmaceuticals has been developed. Its elution characteristics and chemical stability merit its consideration as a replacement for present generators. The 99Mo is present as a zirconium molybdate gel, the high molybdate content of which allows the use of (n, gamma) 99Mo. The resulting generator is as effective and as convenient to use as chromatographic generators that are based on fission product 99Mo. Laboratory evaluations of the concept at low levels of radioactivity described in this paper show that the gel can be prepared in a form which is stable when used in a generator and from which TcO4- can be eluted in yields of 80-85%. The influence of formulation variables on gel properties was established. The performance of this generator using 99Mo with specific activities between 4 and 13 GBq g-1 is described in a further paper.

Zirconium molybdate gel as a generator for technetium-99m--II. High activity generators.

The characteristics of zirconium molybdate gels were described in Part I of this paper. Here it is shown that zirconium molybdate gel, prepared from neutron irradiated molybdenum oxide with 99Mo specific activities between 4.3 and 13.5 GBq g-1 is sufficiently stable for use as a matrix in preparation of high activity 99mTc generators. Elution efficiencies of generators containing from 2 to 6 g of gel ranged from 83 to 50% for 10 mL elutions. Generators containing 150 g of gel were eluted with efficiencies of 80-98%, with the activity peak in the first 50 mL. Radionuclidic impurities were considerably below the limits set by the British Pharmacopoeia. Although the pH of eluates was less than 4, subsequent treatment with a small zirconium oxide bed provided adjustment to within BP limits, and reduced 99Mo impurity levels tenfold to values near 10(-4)%. No significant differences were found between the biodistributions in rats of radiopharmaceuticals prepared with pertechnetate from gel generators and those from other generators.