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.     

The involvement of midbrain astrocyte in the development of morphine tolerance

Aims

Systemic administration of opiate analgesics such as morphine remains the most effective treatment for alleviating severe pain across a range of conditions including acute pain. However, chronic or repeated administration of opiate analgesics results in the development of analgesic tolerance. Glial cells such as microglia and astrocytes are known to release various inflammatory cytokines and neurotrophic factors leading to regulation of neuronal function. Recently, glial cells were reported to play important roles in the development of analgesic tolerance to morphine. Here, we focused on the involvement of midbrain glial cells, particularly astrocytes, in the development of analgesic tolerance to morphine.

Main methods

Mice were treated with morphine (10 mg/kg, s.c.) or vehicle once a day for 5 days. Pentoxifylline (an inhibitor of glial activation; 20 mg/kg, i.p. or 50 and 100 μg/mouse, i.c.v.) was administered 30 min before morphine treatment. Flavopiridol (a cyclin-dependent kinase inhibitor; 5 nmol/mouse, i.c.v.) was administered 10 min before and 10 h after morphine treatment. The analgesic effect of morphine was measured using the tail flick method.

Key findings

The development of analgesic tolerance to morphine was gradually observed during daily treatment of morphine for 5 days in mice. On days 1 and 3 after repeated morphine treatment, astrocyte marker glial fibrillary acidic protein expression levels were significantly increased, as determined by western blot analyses. These phenomena were significantly inhibited following pre-treatment with pentoxifylline or flavopiridol.

Significance

We demonstrated that midbrain astrocytes play an important role in the development of analgesic tolerance to morphine.     

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.     

Representation of lateral line and electrosensory systems in the midbrain of the axolotl,Ambystoma mexicanum

Summary The study focussed on the representation of the electrosensory and lateral line units in the midbrain of the axolotl Ambystoma mexicanum. In addition, the responses to photic and acoustic/vibrational stimuli were determined. Unit properties were characterized with respect to baseline activity, sensitivity, latency, directional specificity and number of input modalities. The anatomical arrangement of the units was determined using stereotactic and histological measurements of the electrode positions.Of 106 units recorded, 29 units were unimodal, 77 units responded to more than one modality. Most units discharged only in response to stimuli. Thresholds of electrosensory units were about 100 V/cm field strength; lateral line units had thresholds below 5 m pp amplitude. The shortest latencies (8–17 ms) were found for responses to visual stimuli. Lateral line and vestibular units responded after 35–58 ms, electroreceptive units after 79–150 ms. All electrosensory and about 50% of the lateral line units were sharply tuned to definite stimulus directions.Electrosensory and lateral line units formed topographical maps in the tectum. The map in each tectal hemisphere contained information about the contralateral surroundings. The electrosensory, lateral line and visual representations were only partly in register; especially in the caudal areas of the midbrain the alignment was poor.     

Evidence for amygdaloid projections to the contralateral hypothalamus and the ipsilateral midbrain in the rat

Summary Light microscopic autoradiography was performed subsequent to injection of tritiated amino acids into various parts of the amygdaloid body of the rat. Evidence is provided for two hitherto unreported projections of the amygdala: from the medial amygdaloid nucleus to the contralateral premamillary nuclei and from the central amygdaloid nucleus to the mesencephalic central grey. The functional implications of these findings are discussed.     

Xenopus axin interacts with glycogen synthase kinase-3 beta and is expressed in the anterior midbrain

Axin is encoded by the fused locus in mice and is required for normal vertebrate axis formation. It has recently been shown that axin associates with the adenomatous polyposis coli gene product (APC), β-catenin and glycogen synthase kinase-3 (GSK-3) in a complex that appears to regulate the level of cytoplasmic β-catenin. We have identified the Xenopus homologue of axin through its interaction with GSK-3β. Xenopus axin (Xaxin) is expressed maternally and throughout early development with a low level of ubiquitous expression. Xaxin also shows remarkably high expression in the anterior mesencephalon adjacent to the forebrain–midbrain boundary.     

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.     

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.