Coding of amplitude modulation in the auditory midbrain of the bullfrog (Rana catesbeiana) across metamorphosis

The functional development of the auditory system across metamorphosis was examined by recording neural activity from the torus semicircularis of larval and postmetamorphic bullfrog froglets in response to amplitude-modulated sound. Multiunit activity in the torus semicircularis during early larval stages showed significant phase-locking to the envelopes of amplitude-modulated noise bursts, up to modulation rates as high as 250 Hz. Beginning at metamorphic climax and continuing into the froglet period, phase locking was restricted to the more limited frequency range characteristic of adult frogs. The onset of operation of the tympanic pathway does not reinstate the highly synchronous neural activity characteristic of the operation of the fenestral pathway. Modulation transfer functions based on spike count did not show tuning for modulation rate in early stage tadpoles, but a greater variety of shapes of these functions emerged as development proceeded. Most of the different kinds of modulation transfer functions seen in adult frogs were also observed in froglets, but band-pass functions were not as sharply peaked. These data suggest that different neural codes for processing of the periodicity of complex signals operate in early stage tadpoles than in postmetamorphic froglets. Accepted: 7 October 1998     

Ontogenetic changes in the auditory sensitivity of damselfishes (pomacentridae)

Psychophysical experiments demonstrated that the hearing abilities of two damselfish species change during ontogeny. Auditory thresholds of four size-groups of juvenile bicolor damselfish. Pomacentrus partitus, and for comparative purposes, three different sized juveniles belonging to a closely related species, P. variablis, were determined through classical conditioning experiments conducted in a standing wave tube. Young juveniles (10–27 days post-metamorphosis) exhibited poor hearing, with detection limens ranging from 54 dB (at 300 Hz) to 15 dB (at 1500 Hz) higher than known adult thresholds. Thresholds decreased exponentially with increasing age, while rapidly approaching adult levels. Youngest juveniles exhibited flat, untuned audiograms, with the appearance of a best frequency and a pattern of increasing acuity progressing in a manner similar to that observed in mammals and birds.     

Female reproductive state influences the auditory midbrain response

Female behavioral responses to sensory stimuli can be highly variable across the reproductive cycle. Female green treefrogs (Hyla cinerea) use the male vocal signal to locate and choose a mate. Gravid females approach a vocalizing male to mate but do not approach if they have recently mated. Such differences in behavioral response may be due in part to shifts in the neural representation of auditory information in the brain. In this study, we investigated the influence of female reproductive state on neural responses in the auditory midbrain to both communication signals (advertisement calls) and non-communication sounds (band limited noise bursts). Recently mated females exhibited significantly reduced response strengths compared to females not recently mated. Reduced response strengths in post-mated females were in response to both noise bursts and male advertisement calls but were limited to the lower frequency range corresponding to the amphibian papilla of the peripheral auditory system. Our results therefore show that the ability of social signals to stimulate the auditory system differs in females depending on their reproductive state, and that the differential effect on low versus high spectral sensitivities may influence the way the two spectral peaks of male advertisement calls are represented.     

Developmental changes in cell proliferation in the auditory midbrain of the bullfrog, Rana catesbeiana

We examined patterns of cell proliferation in the auditory midbrain (torus semicircularis) of the bullfrog, Rana catesbeiana, over larval and early postmetamorphic development, by visualizing incorporation of 5-bromo-2'-deoxyuridine (BrdU) in cycling cells. At all developmental stages, BrdU-labeled cells were concentrated around the optic ventricle. BrdU-labeled cells also appeared within the torus semicircularis itself, in a stage-specific manner. The mitotic index, quantified as the percent of BrdU-positive cells outside the ventricular zone per total cells available for label, varied over larval development. Mitotic index was low in hatchling, early larval, and late larval stages, and increased significantly in deaf period, metamorphic climax, and froglet stages. Cell proliferation was higher in metamorphic climax than at other stages, suggesting increased cell proliferation in preparation for the transition from an aquatic to an amphibious existence. The change in mitotic index over development did not parallel the change in the total numbers of cells available for label. BrdU incorporation was additionally quantified by dot-blot assay, showing that BrdU is available for label up to 72 h postinjection. The pattern of change in cell proliferation in the torus semicircularis differs from that in the auditory medulla (dorsal medullary nucleus and superior olivary nucleus), suggesting that cell proliferation in these distinct auditory nuclei is mediated by different underlying mechanisms.     

Encoding of phase spectra by the peripheral auditory system of the bullfrog

In this study we have examined the sensitivity of auditory nerve fibers in the bullfrog (Rana catesbeiana) to changes in the phase spectrum of an equal-amplitude multi-harmonic stimulus which spanned the bullfrog's range of hearing. To assess peripheral auditory phase sensitivity, changes in the response properties of VIIIth nerve fibers were measured when the relative phase angle of a single harmonic component nearest a unit's best excitatory frequency was systematically varied. The results revealed that shifts in the phase spectrum are encoded in at least J different ways by the peripheral auditory system of the bullfrog: 1) by changes in the degree of spike synchronization of fibers from both inner ear organs (the amphibian papilla and the basilar papilla) to the fundamental waveform period; 2) by changes in the shapes of period histograms of fibers from both organs; and 3) by changes in the spike rates of amphibian papilla fibers. The presence of phase sensitivity in the peripheral auditory system of the bullfrog indicates that information regarding the fine-temporal waveshape and the underlying phase spectrum of an acoustic signal is contained within the spike trains of VIIIth nerve fibers. Similar sensitivities to changes in the phase spectra and temporal waveshapes of acoustic signals may also be present in the peripheral auditory system of other vertebrates. Such studies could provide valuable insight into the role that phase spectra and temporal waveshape may play in bioacoustic communication.Abbreviations BEF best excitatory frequency - BEC best excitatory component - CS_{f 1} synchronization to the fundamental period Portions of this study have been summarized in abstract form (Bodnar and Capranica 1991)     

Postmetamorphic changes in parvalbumin expression in the hindlimb skeletal muscle of the bullfrog,Rana catesbeiana

Anuran amphibians, animals that spend a terrestrial life after metamorphosis, exhibit a marked development of hindlimbs during and after metamorphosis. In order to see whether changes occur in the muscle protein components in the course of postmetamorphic development, we subjected gastrocnemius muscle extracts from growing froglets to two-dimensional electrophoresis (2DE). As a result, we found two proteins to undergo a change in level. One spot, indicating a molecular mass of approximately 12 kDa and an isoelectric point (pI) of 5.0 first became detectable at 45 days after metamorphosis. Another spot, corresponding to a protein of 11 kDa and pI 4.8, was prominent until the former spot appeared. N-terminal amino acid sequence analysis and comparison of the spots with those of parvalbumin (PA) revealed that these two proteins were PA alpha and PA beta. Northern blot analysis using PA alpha and PA beta cDNAs as probes revealed that the PA beta mRNA level declined whereas that of PA alpha mRNA rose as the frogs grew.     

Cell type-specificity of nonclassical estrogen signaling in the developing midbrain

Estrogens have widespread biological functions in the CNS involving the coordination of developmental processes, the regulation of cell physiology, and the control of neuroendocrine systems. In the midbrain, estrogens promote the survival, maturation, and function of neurons and, in particular, of dopamine cells. Aside from classical signaling through nuclear estrogen receptors, we have provided evidence that cellular transmission of estrogen effects in the midbrain comprises a complex intracellular signaling scenario. The major conclusion drawn from our studies is that estrogens interact with yet unidentified membrane receptor complexes which stimulate the phospholipase C and induce the formation of inosite-tri-phosphate (IP₃). This causes a rapid and transitory rise in intracellular free calcium. The modulation of calcium homeostasis is the primary nonclassical physiological response to estrogens in all cell types. Surprisingly, a different secondary downstream signaling cascade seems to be activated in each estrogen-responsive cell population, i.e. phosphatidylinositol-3 kinase (PI3-kinase) in GABAergic and cAMP/ protein kinase A (PKA) in dopaminergic neurons, mitogen-activated protein kinase (MAP-kinase) in astrocytes. The precise biological role of estrogens for the different cell types is still fragmentary. We assume that estrogens positively influence intracellular signaling mechanisms which are important for cell differentiation and survival. It remains to be elucidated what determines the cell type-specificity of these estrogen responses.     

EphB receptor tyrosine kinases control morphological development of the ventral midbrain

EphB receptor tyrosine kinases and ephrin-B ligands regulate several types of cell-cell interactions during brain development, generally by modulating the cytoskeleton. EphB/ephrinB genes are expressed in the developing neural tube of early mouse embryos with distinct overlapping expression in the ventral midbrain. To test EphB function in midbrain development, mouse embryos compound homozygous for mutations in the EphB2 and EphB3 receptor genes were examined for early brain phenotypes. These mutants displayed a morphological defect in the ventral midbrain, specifically an expanded ventral midline evident by embryonic day E9.5-10.5, which formed an abnormal protrusion into the cephalic flexure. The affected area was comprised of cells that normally express EphB2 and ephrin-B3. A truncated EphB2 receptor caused a more severe phenotype than a null mutation, implying a dominant negative effect through interference with EphB forward (intracellular) signaling. In mutant embryos, the overall number, size, and identity of the ventral midbrain cells were unaltered. Therefore, the defect in ventral midline morphology in the EphB2;EphB3 compound mutant embryos appears to be caused by cellular changes that thin the tissue, forcing a protrusion of the ventral midline into the cephalic space. Our data suggests a role for EphB signaling in morphological organization of specific regions of the developing neural tube.     

Hormone-induced vocal behavior and midbrain auditory sensitivity in the green treefrog,Hyla cinerea

Summary Twenty four castrated male, 6 intact male, and 11 intact female Hyla cinerea were injected subcutaneously with 25 g arginine-vasotocin (AVT) and induced to call 1 h later in response to the playback of a conspecific mating call. Eighteen castrated males and 8 intact females were implanted 5 mg androgen pellets for 3 weeks prior to the neuropeptide injection. Among castrated males, 6/9 testosterone (T) implanted, 4/9 dihydrotestosterone (DHT) implanted and 2/6 non implanted individuals produced calls after being administered AVT. 5/6 intact non implanted males and 6/8 T intact implanted females also called, and 3 intact non implanted females remained silent after the injection. Evoked calls had a mid-frequency spectral peak at about 1900 Hz which is absent in field-recorded mating calls of this species. Calls of implanted females and castrated non implanted males were shorter than those of castrated implanted and intact non implanted males. Audiograms measured before hormone implants showed dips of enhanced sensitivity at about 0.5, 0.9 and 3.0 kHz in males and females. After AVT injection, thresholds at frequencies within the 0.7–1.5 kHz range were increased in castrated males. Such reduction in sensitivity points to an inhibition of the auditory system during hormone induced vocal activation.Abbreviations AVT arginine-vasotocin - DHT dihydrotestosterone - T testosterone - TS torus semicircularis     

Basic maps in the auditory midbrain

Topologic maps at consecutive levels of sensory pathways indicate behaviorally relevant features of stimuli at increasing degrees of complexity. In the auditory system, except for tonotopic maps, the nature of represented features is unknown. In a model analogous to visual map formation we show that in the auditory midbrain, layers of neurons with preference to the same frequency (isofrequency planes) may hold maps of two basic, mutually orthogonal parameters--instantaneous amplitude and phase--of basilar membrane displacement at the cochlear location responding to that frequency. The proposed neural tuning to frequency, amplitude, and phase implies that sound is transformed into specific temporal trajectories of neural activation, with consequences for experimental design and interpretation of neural response behavior.     

Metamorphosis-related changes in the lateral line system of lampreys, <Emphasis Type="Italic">Petromyzon marinus</Emphasis>

Lamprey metamorphosis leads to considerable changes in morphology and behavior. We have recently reported that larval lampreys possess a functional lateral line system. Here we investigated metamorphic morphological changes in the lateral line system using light and electron microscopy. Functional modifications were studied by recording the trunk lateral line nerve activity of larvae and adults while stimulating neuromasts with approximately sinusoidal water motion. We found a general re-patterning of neuromasts on the head and trunk including an increase in numbers, redistribution within the pit lines, and shifts of the pit lines relative to external features. The trunk lateral line nerve response was qualitatively similar in adults and larvae. Both showed two neuronal populations responding to opposite directions of water flow. Magnitude of the response increased monotonically with stimulus amplitude. At low frequencies, the response lag relative to the stimulus maximum was approximately 220°, and the gain depended approximately linearly on frequency, confirming that superficial neuromasts are velocity detectors. Changes in phase lag with increasing stimulus frequency were steeper in larvae, suggesting slower afferent conductance. The response gain with frequency was smaller for adults, suggesting a narrower frequency discrimination range and decreased sensitivity. These changes may be adaptations for the active lifestyle of adult lampreys.     

Coding of lateral line stimuli in the goldfish midbrain in still and running water

We investigated in goldfish, Carassius auratus, how running water affects the responses of toral lateral line units to a stationary vibrating sphere or to a non-vibrating sphere that moves along the side of the fish. Experiments were conducted in the presence of running water (hydrodynamic noise) to further explore the sensory capabilities of the lateral line with special focus on the morphological sub-modalities. Previous recordings from lateral line nerve fibres in various fish species and the first nucleus of the ascending lateral line pathway in goldfish revealed flow-sensitive and flow-insensitive units. These physiological differences represent, at least in part, the differences in morphology of the lateral line, superficial and canal neuromasts. Following up on these findings we recorded flow-sensitive and flow-insensitive units in the Torus semicircularis of goldfish. In still water, both types of units responded to a vibrating or moving sphere. In running water, neural responses were weaker when the sphere was moved with the flow but were comparable or slightly stronger when the sphere was moved against the flow. In running water, responses of flow-sensitive fibres to the vibrating sphere were masked. In contrast, the responses of units insensitive to water flow were not masked. Our data confirm previous findings but also indicate differences when compared to previous reports. We discuss these differences with respect to lateral line morphology, sub-modalities and convergence of different channels of information at higher brain stations.     

Interactions of bullfrog tadpole predators and an insecticide: predation release and facilitation

The effect of a contaminant on a community may not be easily predicted, given that complex changes in food resources and predator-prey dynamics may result. The objectives of our study were to determine the interactive effects of the insecticide carbaryl and predators on body size, development, survival, and activity of tadpoles of the bullfrog (Rana catesbeiana). We conducted the study in cattle tank mesocosm ponds exposed to 0, 3.5, or 7.0 mg/l carbaryl, and no predators or two red-spotted newts (Notophthalmus viridescens), bluegill sunfish (Lepomis macrochirus), or crayfish (Orconectes sp.). Carbaryl negatively affected predator survival by eliminating crayfish from all ponds, and by eliminating bluegill sunfish from ponds exposed to the highest concentration of carbaryl; carbaryl exposure did not effect survival of red-spotted newts. Because crayfish were eliminated by carbaryl, bullfrogs were released from predation and survival was near that of predator controls at low concentrations of carbaryl exposure. High concentrations of carbaryl reduced tadpole survival regardless of whether predators survived carbaryl exposure or not. Presence of crayfish and newts reduced tadpole survival, while bluegill sunfish appeared to facilitate bullfrog tadpole survival. Presence of carbaryl stimulated bullfrog tadpole mass and development. Our study demonstrates that the presence of a contaminant stress can alter community regulation by releasing prey from predators that are vulnerable to contaminants in some exposure scenarios.Due to an error in the citation line, this revised PDF (published in December 2003) deviates from the printed version, and is the correct and authoritative version of the paper.     

Seasonal changes in frequency tuning and temporal processing in single neurons in the frog auditory midbrain

Frogs rely on acoustic signaling to detect, discriminate, and localize mates. In the temperate zone, reproduction occurs in the spring, when frogs emerge from hibernation and engage in acoustically guided behaviors. In response to the species mating call, males typically show evoked vocal responses or other territorial behaviors, and females show phonotactic responses. Because of their strong seasonal behavior, it is possible that the frog auditory system also displays seasonal variation, as evidenced in their vocal control system. This hypothesis was tested in male Northern leopard frogs by evaluating the response characteristics of single neurons in the torus semicircularis (TS; a homolog of the inferior colliculus) to a synthetic mating call at different times of the year. We found that TS neurons displayed a seasonal change in frequency tuning and temporal properties. Frequency tuning shifted from a predominance of TS units sensitive to intermediate frequencies (700-1200 Hz) in the winter, to low frequencies (100-600 Hz) in the summer. In winter and early spring, most TS neurons showed poor, or weak, time locking to the envelope of the amplitude-modulated synthetic call, whereas in late spring and early summer the majority of TS neurons showed robust time-locked responses. These seasonal differences indicate that neural coding by auditory midbrain neurons in the Northern leopard frog is subject to seasonal fluctuation.     

Catecholamines induce metamorphosis in the hydrozoan Halocordyle disticha but not in Hydractinia echinata

Summary Planula larvae of the marine hydroids Halocordyle disticha and Hydractinia echinata were treated with the catecholamines epinephrine, norepinephrine and dopamine, as well as with certain of their precursors and agonists. Norepinephrine, l-dopa, dopamine and the dopamine agonist ADTN at concentrations ranging from 0.1 to 0.001 mM induced metamorphosis within 24 h in Halocordyle disticha, with no observable morphogenetic abnormalities. Epinephrine, the adrenergic agonists phenylephrine, isoproterenol and methoxyamine, and the catecholamine precursors phenylalanine and tyrosine were found not to induce metamorphosis at the concentrations employed. None of the compounds was effective in inducing metamorphosis in Hydractinia echinata. A model is presented for neural control of metamorphosis in Halocordyle disticha     

Developmental and diel profiles of plasma corticosteroids in the bullfrog,Rana catesbeiana

Corticosteroids synergize with the thyroid hormone (TH) at late metamorphic stages and might have a role in the hormonal regulation of amphibian metamorphosis. This role could be influenced by diel fluctuations, particularly if the peak of the plasma corticoids changed in relation to the TH peaks. Diel variation in plasma corticosteroids was studied in Rana catesbeiana prometamorphic and climax tadpoles on 18:6, 12:12 and 6:18 light:dark (LD) cycles. Cortisol (hydrocortisone; HC) and aldosterone (ALDO) exhibited different, but LD cycle-specific, circadian fluctuations at prometamorphosis, whereas corticosterone (CORT) was undetectable (less than 1.18 ng/ml). HC, ALDO and CORT rhythms became synchronous at early metamorphic climax on all LD cycles, although the cosinor-derived acrophases, which occurred around the time of the dark:light transition, shifted approximately 6 h earlier from 18L:6D to 6L:18D. On both 18L:6D and 12L:12D, the acrophase of HC changed little from prometamorphosis to climax, whereas that of ALDO underwent a major phase shift. On 6L:18D, both the ALDO and the HC acrophases shifted at climax. These LD cycle-specific phase shifts of the diel rhythms placed the acrophases of the corticoids in different phase relationships to that of the previously determined thyroxine (T(4)) acrophase at climax, and may partially explain the influence of the light regimen on metamorphic timing. The pronounced diel variations in the corticoid concentrations from the troughs to the peaks show that hormone levels are a function of the time of day and the environmental lighting regimen, which need to be taken into account in measuring the level of plasma hormones in amphibians. The 24-h means calculated from the data of all the sampling times showed that only plasma ALDO and CORT, but not HC, rose markedly at climax, although there were significant LD cycle-related differences in the mean levels of both HC and ALDO at prometamorphosis, and in HC at climax. Additional work sampling at mid-light showed that plasma CORT peaked at Stage XXIII, decreased at the end of climax, and remained low in the postmetamorphic froglet at 2.1 ng/ml. In the adult bullfrog, CORT was clearly the predominant corticosteroid at 34.3 ng/ml, whereas HC and ALDO levels were only approximately 1.3 ng/ml.     

Micro-computed tomography of pupal metamorphosis in the solitary bee Megachile rotundata

Insect metamorphosis involves a complex change in form and function. In this study, we examined the development of the solitary bee, Megachile rotundata, using micro-computed tomography (μCT) and volume analysis. We describe volumetric changes of brain, tracheae, flight muscles, gut, and fat bodies in prepupal, pupal, and adult M. rotundata. We observed that individual organ systems have distinct patterns of developmental progression, which vary in their timing and duration. This has important implications for commercial management of this agriculturally relevant pollinator.