How frequency hopping suppresses pulse-echo ambiguity in bat biosonar.

Big brown bats transmit wideband FM biosonar sounds that sweep from 55 to 25 kHz (first harmonic, FM1) and from 110 to 50 kHz (second harmonic, FM2). FM1 is required to perceive echo delay for target ranging; FM2 contributes only if corresponding FM1 frequencies are present. We show that echoes need only the lowest FM1 broadcast frequencies of 25 to 30 kHz for delay perception. If these frequencies are removed, no delay is perceived. Bats begin echo processing at the lowest frequencies and accumulate perceptual acuity over successively higher frequencies, but they cannot proceed without the low-frequency starting point in their broadcasts. This reveals a solution to pulse-echo ambiguity, a serious problem for radar or sonar. In dense, extended biosonar scenes, bats have to emit sounds rapidly to avoid collisions with near objects. But if a new broadcast is emitted when echoes of the previous broadcast still are arriving, echoes from both broadcasts intermingle, creating ambiguity about which echo corresponds to which broadcast. Frequency hopping by several kilohertz from one broadcast to the next can segregate overlapping narrowband echo streams, but wideband FM echoes ordinarily do not segregate because their spectra still overlap. By starting echo processing at the lowest frequencies in frequency-hopped broadcasts, echoes of the higher hopped broadcast are prevented from being accepted by lower hopped broadcasts, and ambiguity is avoided. The bat-inspired spectrogram correlation and transformation (SCAT) model also begins at the lowest frequencies; echoes that lack them are eliminated from processing of delay and no longer cause ambiguity.

Cholesterol selectively regulates IL-5 induced mitogen activated protein kinase signaling in human eosinophils.

Eosinophils function contributes to human allergic and autoimmune diseases, many of which currently lack curative treatment. Development of more effective treatments for eosinophil-related diseases requires expanded understanding of eosinophil signaling and biology. Cell signaling requires integration of extracellular signals with intracellular responses, and is organized in part by cholesterol rich membrane microdomains (CRMMs), commonly referred to as lipid rafts. Formation of these organizational membrane domains is in turn dependent upon the amount of available cholesterol, which can fluctuate widely with a variety of disease states. We tested the hypothesis that manipulating membrane cholesterol content in primary human peripheral blood eosinophils (PBEos) would selectively alter signaling pathways that depend upon membrane-anchored signaling proteins localized within CRMMs (e.g., mitogen activated protein kinase [MAPK] pathway), while not affecting pathways that signal through soluble proteins, like the Janus Kinase/Signal Transducer and Activator of Transcription [JAK/STAT] pathway. Cholesterol levels were increased or decreased utilizing cholesterol-chelating methyl-ß-cyclodextrin (MßCD), which can either extract membrane cholesterol or add exogenous membrane cholesterol depending on whether MßCD is preloaded with cholesterol. Human PBEos were pretreated with MßCD (cholesterol removal) or MßCD+Cholesterol (MßCD+Chol; cholesterol delivery); subsequent IL-5-stimulated signaling and physiological endpoints were assessed. MßCD reduced membrane cholesterol in PBEos, and attenuated an IL-5-stimulated p38 and extracellular-regulated kinase 1/2 phosphorylation (p-p38, p-ERK1/2), and an IL-5-dependent increase in interleukin-1ß (IL-1ß) mRNA levels. In contrast, MßCD+Chol treatment elevated PBEos membrane cholesterol levels and basal p-p38, but did not alter IL-5-stimulated phosphorylation of ERK1/2, STAT5, or STAT3. Furthermore, MßCD+Chol pretreatment attenuated an IL-5-induced increase in cell survival at 48 hours, measured as total cellular metabolism. The reduction in cell survival following cholesterol addition despite unaltered STAT phosphorylation contradicts the current dogma in which JAK/STAT activation is sufficient to promote eosinophil survival, and suggests an additional, unidentified mechanism critically regulates IL-5-mediated human PBEos survival.

Spatial release from simultaneous echo masking in bat sonar.

Big brown bats (Eptesicus fuscus) use biosonar to navigate and locate objects in their surroundings. During natural foraging, they often encounter echoes returned by a target of interest located to the front while other, often stronger, clutter echoes are returned from objects, such as vegetation, located to the sides or above. Nevertheless, bats behave as if they do not suffer interference from this clutter. Using a two-choice delay discrimination procedure, bats were tested for the masking effectiveness of clutter echoes on target echoes when the target echoes were delivered from the bat's front while clutter echoes were delivered from 90° overhead, a direction of lowpass filtering by the external ears. When clutter echoes are presented from the front at the same delay as target echoes, detection performance declines and clutter masking occurs. When the clutter echoes are presented at the same delay but from overhead, discrimination performance is unaffected and no masking occurs. Thus there is masking release for simultaneous off-axis lowpass clutter compared to masking by simultaneous clutter from the front. The bat's performance for simultaneous target and clutter echoes indicates a new role for the mechanism that separates overlapping echoes by decomposing the bat's auditory time-frequency representation.

Human airway eosinophils exhibit preferential reduction in STAT signaling capacity and increased CISH expression.

Allergic asthma, a chronic respiratory disorder marked by inflammation and recurrent airflow obstruction, is associated with elevated levels of IL-5 family cytokines and elevated numbers of eosinophils (EOS). IL-5 family cytokines elongate peripheral blood EOS (EOS(PB)) viability, recruit EOS(PB) to the airways, and, at higher concentrations, induce degranulation and reactive oxygen species generation. Although airway EOS (EOS(A)) remain signal ready in that GM-CSF treatment induces degranulation, treatment of EOS(A) with IL-5 family cytokines no longer confers a survival advantage. Because the IL-5 family receptors have common signaling capacity, but are uncoupled from EOS(A) survival, whereas other IL-5 family induced endpoints remain functional, we tested the hypothesis that EOS(A) possess a JAK/STAT-specific regulatory mechanism (because JAK/STAT signaling is critical to EOS survival). We found that IL-5 family-induced STAT3 and STAT5 phosphorylation is attenuated in EOS(A) relative to blood EOS from airway allergen-challenged donors. However, IL-5 family-induced ERK1/2 phosphorylation is not altered between EOS(A) and EOS from airway allergen-challenged donors. These observations suggest EOS(A) possess a regulatory mechanism for suppressing STAT signaling distinct from ERK1/2 activation. Furthermore, we found, in EOS(PB), IL-5 family cytokines induce members of the suppressors of cytokine signaling (SOCS) genes, CISH and SOCS1. Additionally, following allergen challenge, EOS(A) express significantly more CISH and SOCS1 mRNA and CISH protein than EOS(PB) counterparts. In EOS(PB), long-term pretreatment with IL-5 family cytokines, to varying degrees, attenuates IL-5 family-induced STAT5 phosphorylation. These data support a model in which IL-5 family cytokines trigger a selective downregulation mechanism in EOS(A) for JAK/STAT pathways.

Bats use echo harmonic structure to distinguish their targets from background clutter.

When echolocating big brown bats fly in complex surroundings, echoes arriving from irrelevant objects (clutter) located to the sides of their sonar beam can mask perception of relevant objects located to the front (targets), causing "blind spots." Because the second harmonic is beamed more weakly to the sides than the first harmonic, these clutter echoes have a weaker second harmonic. In psychophysical experiments, we found that electronically misaligning first and second harmonics in echoes (to mimic the misalignment of corresponding neural responses to harmonics in clutter echoes) disrupts the bat's echo-delay perception but also prevents clutter masking. Electronically offsetting harmonics to realign their neural responses restores delay perception but also clutter interference. Thus, bats exploit harmonics to distinguish clutter echoes from target echoes, sacrificing delay acuity to suppress masking.

Perception of echo delay is disrupted by small temporal misalignment of echo harmonics in bat sonar.

Echolocating big brown bats emit ultrasonic frequency-modulated (FM) biosonar sounds containing two prominent downward-sweeping harmonics (FM1 and FM2) and perceive target distance from echo delay. In naturally occurring echoes, FM1 and FM2 are delayed by the same amount. Even though echoes from targets located off-axis or far away are lowpass filtered, which weakens FM2 relative to FM1, their delays remain the same. We show here that misalignment of FM2 with FM1 by only 2.6 µs is sufficient to significantly disrupt acuity, which then persists for larger misalignments up to 300 µs. However, when FM2 is eliminated entirely rather than just misaligned, acuity is effectively restored. For naturally occurring, lowpass-filtered echoes, neuronal responses to weakened FM2 are retarded relative to FM1 because of amplitude-latency trading, which misaligns the harmonics in the bat's internal auditory representations. Electronically delaying FM2 relative to FM1 mimics the retarded neuronal responses for FM2 relative to FM1 caused by amplitude-latency trading. Echoes with either electronically or physiologically misaligned harmonics are not perceived as having a clearly defined delay. This virtual collapse of delay acuity may suppress interference from off-axis or distant clutter through degradation of delay images for clutter in contrast to sharp images for nearer, frontal targets.

Effects of filtering of harmonics from biosonar echoes on delay acuity by big brown bats (Eptesicus fuscus).

Big brown bats emit FM biosonar sounds containing two principal harmonics (FM1 approximately 55-22 kHz;FM2 approximately 105-45 kHz). To examine the role of harmonics, they were selectively filtered from stimuli in electronic-echo delay discrimination experiments. Positive stimuli were delayed by 3.16 ms (55 cm simulated target range); negative stimuli were by delayed by 3.96 ms (68 cm). This large 800-micros delay difference (nearly 14 cm) was easily discriminated for echoes containing equal-strength FM1 and FM2. Performance gradually decreased as highpass filters removed progressively larger segments from FM1. For echoes with FM2 alone, performance collapsed to chance, but performance remained good for lowpass echoes containing FM1 alone. Attenuation of FM2 by 3 dB relative to FM1 also decreased performance, but shortening electronic delay of the attenuated FM2 by 48 micros counteracted amplitude-latency trading and restored performance. Bats require the auditory representations of FM1 and FM2 to be in temporal register for high delay acuity. Misalignment of neuronal responses degrades acuity, but outright removal of FM2, leaving only FM1, causes little loss of acuity. Functional asymmetry of harmonics reflects lowpass effects from beaming and atmospheric propagation, which leave FM1 intact. It may cooperate with latency shifts to aid in suppression of clutter.

Spatial location influences vocal interactions in bullfrog choruses.

A multiple sensor array was employed to identify the spatial locations of all vocalizing male bullfrogs (Rana catesbeiana) in five natural choruses. Patterns of vocal activity collected with this array were compared with computer simulations of chorus activity. Bullfrogs were not randomly spaced within choruses, but tended to cluster into closely spaced groups of two to five vocalizing males. There were nonrandom, differing patterns of vocal interactions within clusters of closely spaced males and between different clusters. Bullfrogs located within the same cluster tended to overlap or alternate call notes with two or more other males in that cluster. These near-simultaneous calling bouts produced advertisement calls with more pronounced amplitude modulation than occurred in nonoverlapping notes or calls. Bullfrogs located in different clusters more often alternated entire calls or overlapped only small segments of their calls. They also tended to respond sequentially to calls of their farther neighbors compared to their nearer neighbors. Results of computational analyses showed that the observed patterns of vocal interactions were significantly different than expected based on random activity. The use of a multiple sensor array provides a richer view of the dynamics of choruses than available based on single microphone techniques.

FM echolocating bats shift frequencies to avoid broadcast-echo ambiguity in clutter.

Sonar broadcasts are followed by echoes at different delays from objects at different distances. When broadcasts are emitted rapidly in cluttered surroundings, echo streams from successive broadcasts overlap and cause ambiguity in matching echoes to corresponding broadcasts. To identify reactions to ambiguity in clutter, echolocating bats that emit multiple-harmonic FM sounds were trained to fly into a dense, extended array of obstacles (multiple rows of vertically hanging chains) while the sonar sounds the bat emitted were recorded with a miniature radio microphone carried by the bat. Flight paths were reconstructed from thermal-infrared video recordings. Successive rows of chains extended more than 6 m in depth, so each broadcast was followed by a series of echoes from multiple rows of chains that lasted up to 40 ms. Bats emitted sounds in pairs ("strobe groups") at short (20-40 ms) interpulse intervals (IPIs) alternating with longer IPIs (>50 ms). For many short IPIs, the stream of echoes from the first broadcast was still arriving when the second broadcast was emitted. This overlap caused ambiguity about matching echoes with broadcasts. Bats shifted frequencies of the first sound in each strobe group upward and the second sound downward by 3-6 kHz. When overlap and ambiguity ceased, frequency shifts ceased also. Frequency differences were small compared with the total broadcast band, which was 75-80 kHz wide, but the harmonic structure of echoes enhances the differences in spectrograms. Bats could use time-frequency comparisons of echoes with broadcasts to assign echoes to the corresponding broadcasts and thus avoid ambiguity.

Role of broadcast harmonics in echo delay perception by big brown bats.

Big brown bats (Eptesicus fuscus) emit frequency-modulated (FM) echolocation sounds containing two principal down-sweeping harmonics (FM(1) approximately 55-25 kHz, FM(2) approximately 105-50 kHz). To determine whether each harmonic contributes to perception of echo delay, bats were trained to discriminate between "split-harmonic" echoes that differed in delay. The bat's broadcasts were picked up with microphones, and FM(1) and FM(2) were separated with highpass and lowpass filters at about 55 kHz, where they overlap in frequency. Both harmonics then were delivered from loudspeakers as positive stimuli in a 2-choice delay discrimination procedure with FM(1) delayed 3.16 ms and FM(2) delayed 3.46 ms (300 mus delay split). Negative stimuli contained FM(1) and FM(2) with the same filtering but no delay separation. These were presented at different overall delays from 11 down to 3 ms to measure the bat's delay discrimination acuity for each harmonic in the split harmonic echoes. The bats determined the delays of both FM(1) and FM(2), but performance was overlaid by a broad pedestal of poor performance that extended for 800 micros. Splitting the harmonics by 300 micros appears to defocus the bat's representation of delay, revealing the existence of a process for recognizing the normally simultaneous occurrence of the harmonics.

Analyzing acoustic interactions in natural bullfrog (Rana catesbeiana) choruses.

Analysis of acoustic interactions between animals in active choruses is complex because of the large numbers of individuals present, their high calling rates, and the considerable numbers of vocalizations that either overlap or show close temporal alternation. The authors describe a methodology for recording chorus activity in bullfrogs (Rana catesbeiana) using multiple, closely spaced acoustic sensors that provide simultaneous estimates of sound direction and sound characteristics. This method provides estimates of location of individual callers, even under conditions of call overlap. This is a useful technique for understanding the complexity of the acoustic scene faced by animals vocalizing in groups.

Jamming avoidance response of big brown bats in target detection.

When searching for prey, big brown bats (Eptesicus fuscus) enhance the range of their sonar by concentrating more energy in the nearly constant-frequency (CF) tail portion of their frequency-modulated (FM) sweeps. We hypothesize that this portion of their signals may be vulnerable to interference from conspecifics using the same frequencies in their own emissions. To determine how bats modify their signals when confronted with an interfering stimulus, we compared the echolocation calls of bats when a CF jamming tone was on and off. The bats performed a two-alternative forced-choice detection task in the laboratory that required the use of echolocation. All three bats shifted the tail-end CF component of their emitted frequency bidirectionally away from the CF jamming stimulus only when the jamming frequency was within 2-3 kHz of the preferred baseline frequency of the bat. The duration of their emissions did not differ between the jamming and no-jamming trials. The jamming avoidance response of bats may serve to avoid masking or interference in a narrow range of frequencies important for target detection.

Expression of interleukin-5- and granulocyte macrophage-colony-stimulating factor-responsive genes in blood and airway eosinophils.

Because interleukin (IL)-5 family cytokines are critical regulators of eosinophil development, recruitment, and activation, this study was initiated to identify proteins induced by these cytokines in eosinophils. Using oligonucleotide microarrays, numerous transcripts were identified as responsive to both IL-5 and granulocyte macrophage-colony-stimulating factor (GM-CSF), but no transcripts were markedly affected by one cytokine and not the other. Expression of several gene products were seen to be increased following in vitro stimulation of human blood eosinophils, including the IL-3 receptor alpha subunit, lymphotoxin beta, Pim-1, and cyclin D3. Given that eosinophils recovered from the bronchoalveolar lavage fluid of allergic patients after antigen challenge are exposed to IL-5 or GM-CSF in the airway prior to isolation, the hypothesis was tested that selected IL-5- and GM-CSF-responsive genes are upregulated in airway eosinophils relative to the expression in blood cells. Airway eosinophils displayed greater cell surface expression of the IL-3 receptor alpha subunit, CD44, CD25, and CD66e, suggesting that these proteins may be markers of eosinophil activation by IL-5 family cytokines in airway eosinophils. Other genes that were induced by both IL-5 and GM-CSF showed protein expression at similar or decreased levels in airway eosinophils relative to their circulating counterparts (i.e., lymphotoxin beta and CD24). These studies have identified several transcriptional targets of IL-5 and GM-CSF in human eosinophils and suggest that a number of protein products are critical to the responsiveness of airway eosinophils.