John A. Thorson

Auditory behavior of the cricket: II. Simplicity of calling-song recognition inGryllus, and anomalous phonotaxis at abnormal carrier frequencies

Summary1.Essential and nonessential features of the male field-cricket (Gryllus campestris L.) calling song have been identified by varying acoustic parameters while testing for phonotactic tracking by females walking on the Kramer spherical treadmill.2.In the natural calling song, a 5-kHz carrier is modulated into 4-syllable chirps, repeated 2–4 times per second; the syllable rate within each chirp is near 30 Hz. Surprisingly, however, the majority of females that are ‘good trackers’ of calling song also track pure ‘trill’ — continuous 30-Hz trains of syllables having no chirp structure — for many minutes. Trill having identical sound power but the wrong syllable rate is not tracked.3.The ratio of syllable duration to syllable repetition interval — that is, the ‘duty cycle’, which is near 50% in nature — can be increased to as much as ca. 90% (intersyllable pauses as brief as 4–5 ms) without degrading tracking performance, as long as syllable repetition rate is near 30 Hz. Similarly, very short duty cycles — in the extreme case songs with syllables consisting of 200-μs ‘clicks’ — can elicit tracking of the sound source by the female.4.Fine details of the temporal structure of the syllables evidently do not convey information in our conditions, because (i)G. campestris females track both the maleG. campestris and the somewhat similar maleG. bimaculatus calling song, and (ii) tracking ofcampestris calling song is indistinguishable from tracking of the same song played backward.5.All the above results are compatible with the view that repetitive modulation near 30 Hz is the chief — and in some cases both necessary and sufficient — parameter of songs that trigger the neuronal ‘recognition’ of male song. Properties of alternative candidate mechanisms for such a process are described.6.As one test that may distinguish among such mechanisms, the range of syllable rates over which recognition is elicited is measured by a paradigm that keeps chirp duration, chirp rate and chirp energy approximately constant. At 80 dB, tracking is observed reliably in the range 20–40 Hz, whereas syllable rates below 15 Hz and above 50 Hz are largely rejected. Multiple-peak structure within this response spectrum, predicted by certain of the recognition mechanisms considered, is — if present — not resolved by our methods.7.Variation of the song parameter ‘carrier frequency’ to values above the natural ca. 5 kHz results in a striking phenomenon at high sound intensities. Maintained ‘tracking’ is observed, but at characteristic, erroneous angles with respect to the sound source. To compare this ‘anomalous phonotaxis’ with the negative phonotaxis reported in other gryllids during flight, we repeat Moiseff et al.s (1978) ‘one-eared circling’ experiments on the Kramer treadmill, in three species of gryllids. InTeleogryllus, walking shows parallels to flight (reversal of one-eared circling in comparisons of 5 and 15 kHz) with respect to the carrier-frequency dependence of phonotaxis. WalkingGryllus andTeleogryllus evidently differ in this respect;Gryllus rarely exhibits negative phonotaxis (by one-ear criteria) at frequencies eliciting anomalous phonotaxis.Finally, we show that simple pressure-gradient effects associated with the ipsilateral spiracular pathways could in principle account for the directional hearing observed inGryllus, and that the same formulation predicts anomalous directions of tracking, qualitatively like those measured, at elevated carrier frequencies. Such trial explanations evidently require drastic modification, however, if they are to account for the behavioral effects of blocking single tympanal and spiracular inputs.

Auditory behavior of the cricket. I: Dynamics of compensated walking and discrimination paradigms on the Kramer treadmill

Summary1.In applying the recently developed methods of compensated walking to the study of cricket phonotaxis, we have first asked how such devices alter the dynamic body-substrate situation from that experienced in natural walking on a fixed substrate. Because a quantitative treatment has not been available, we derive certain general mechanical requirements of such compensation systems, and show in this context just what the Kramer treadmill accomplishes.2.Mathematical predictions of the above analysis were confirmed by frame-by-frame TV analysis of crickets during natural walking and compensated walking on the Kramer treadmill. These are as follows. Body-substrate inertial effects can be made to resemble those occurring during fixed-substrate walking as long as the animal walks continuously and the compensation is highly damped. During starts and stops, however, body-substrate acceleration, and hence the associated leg forces, differ severely from those experienced in walking on a fixed substrate.3.Despite the above effects, the presence of some bearing noise, and the elimination of all visual guidance, we show that a useful form of ‘blind auditory tracking’ can be obtained regularly. Loudspeakers playing the male calling song are accurately tracked by females, with meandering of direction about the target direction, via several characteristic modes of start-stop behavior. The tracking mode adopted at any instant can depend upon song attractiveness, song intensity, the history of stimuli presented, the individual, and factors unknown to us. The mode adopted also affects the nature of the meandering seen in the direction record.4.Depending upon the season and the source of the crickets, one-quarter to one-half of adult females older than 3 to 4 weeks turn out to be ‘good trackers’ in our conditions, and to remain so for some weeks. Sound intensity has little effect upon tracking accuracy in the range 70–90 dB, as long as the song is an attractive one.5.We have developed experimental paradigms which test for discrimination by the female between male calling song and various ‘wrong songs’. These employ two alternating loudspeakers to control against directional clues available from the apparatus, and allow comparison of response to different songs both sequentially and simultaneously (the choice situation). We show that in both kinds of presentation females can clearly and reproducibly discriminate between calling song and wrong songs having no syllable structure or the wrong syllable repetition rate.6.If during the tracking of calling song the song is turned off, the female stops within a few seconds. If, instead, one replaces the calling song suddenly with a wrong song (rejected when presented alone), the wrong song is ‘tracked’ for 10 to 20 s or more. These effects have implications regarding operational definitions of ‘motivation’ and ‘context setting’; they may also lead to ways of distinguishing between the recognition and tracking processes.7.Contrary to expectation based on earlier work withGryllus campestris, we find that increasing the number of syllables per chirp in the range 4–8 (with repetition interval 30 ms) increases, rather than decreases, attractiveness of the song in both our sequential and our choice tests.8.The finding that attractiveness of a song increases with number of syllables suggests that inGryllus the key feature of sound signals that elicit ‘recognition and pursuit’ may be modulation power near 30 Hz, associated with the syllable repetition rate.This hypothesis predicts that natural ‘rivalry song’ (with chirps of 20 or more syllables near the 30-Hz rate) ought to be very attractive; we find that it is, and suggest a reinterpretation of its role in nature.9.In a separate set of experiments, the callingsong sound was gated by the walking-velocity signal so that sound could be heard either (i) only during walking or (ii) only during stops. Only our two best trackers succeeded in our tracking tests if sound was presented strictly during walking; 8 good trackers failed. With sound available only during stops, many succeeded if they happened to employ a suitable tracking mode. Our impression is that ‘stopping to listen’ (or even to look) is ordinarily a significant part of the search behavior.

14-3-3 Proteins Are Required for Maintenance of Raf-1 Phosphorylation and Kinase Activity

ABSTRACT By binding to serine-phosphorylated proteins, 14-3-3 proteins function as effectors of serine phosphorylation. The exact mechanism of their action is, however, still largely unknown. Here we demonstrate a requirement for 14-3-3 for Raf-1 kinase activity and phosphorylation. Expression of dominant negative forms of 14-3-3 resulted in the loss of a critical Raf-1 phosphorylation, while overexpression of 14-3-3 resulted in enhanced phosphorylation of this site. 14-3-3 levels, therefore, regulate the stoichiometry of Raf-1 phosphorylation and its potential activity in the cell. Phosphorylation of Raf-1, however, was insufficient by itself for kinase activity. Removal of 14-3-3 from phosphorylated Raf abrogated kinase activity, whereas addition of 14-3-3 restored it. This supports a paradigm in which the effects of phosphorylation on serine as well as tyrosine residues are mediated by inducible protein-protein interactions.

The 14-3-3 proteins positively regulate rapamycin-sensitive signaling

BACKGROUND The kinase Tor is the target of the immunosuppressive drug rapamycin and is a member of the phosphatidylinositol kinase (PIK)-related kinase family. It plays an essential role in progression through the G1 phase of the cell cycle. The molecular details of Tor signaling remain obscure, however. RESULTS We isolated two Saccharomyces cerevisiae genes, BMH1 and BMH2, as multicopy suppressors of the growth-inhibitory phenotype caused by rapamycin in budding yeast. BMH1 and BMH2 encode homologs of the 14-3-3 signal transduction proteins. Deletion of one or both BMH genes caused hypersensitivity to rapamycin in a manner that was dependent on gene dosage. In addition, alterations in the phosphopeptide-binding pocket of the 14-3-3 proteins had dramatically different effects on their ability to relieve the growth-arresting rapamycin phenotype. Mutations that prevented 14-3-3 from binding to a phosphoserine motif abolished its ability to confer rapamycin resistance. In contrast, substitution of two residues in 14-3-3 that surround these phosphoserine-binding sites conferred a dominant rapamycin-resistant phenotype. CONCLUSIONS Our studies reveal 14-3-3 as an important component in rapamycin-sensitive signaling and provide significant new insights into the structure and function of 14-3-3 proteins.

Lymphocytic Mastitis and Diabetic Mastopathy: A Molecular, Immunophenotypic, and Clinicopathologic Evaluation of 11 Cases

Lymphocytic mastitis and diabetic mastopathy are uncommon fibroinflammatory breast diseases. The lesions seen in these entities are unique in that the associated lymphoid infiltrates are composed of predominantly B cells. In addition, B-cell lymphoepithelial lesions, a finding commonly associated with extranodal marginal zone B-cell/mucosa-associated lymphoid tissue (MALT) lymphomas, are also often present in lymphocytic mastitis and diabetic mastopathy. Although the clinical and immunomorphologic features are well characterized, the clonality of the B-cell infiltrate and the lymphomatous potential of lymphocytic mastitis and diabetic mastopathy have not been emphasized in the literature. We evaluated 11 cases of lymphocytic mastitis/diabetic mastopathy for immunoglobulin heavy chain gene rearrangement and correlated the findings with all available clinical data. A longstanding history of Type I diabetes mellitus was present in seven patients. One nondiabetic patient had Sjogren’s syndrome, and two patients had no history of diabetes mellitus or other autoimmune disease. Clinical data were unavailable for one patient. B-cell–predominant lymphoid infiltrates were seen in all cases, and B-cell lymphoepithelial lesions were found in five. No evidence of a B-cell clone was found in any of the 11 cases by appropriately controlled immunoglobulin heavy chain gene rearrangement studies, and none of the patients developed lymphoma during follow-up intervals ranging from 2–126 months. These findings suggest that despite the presence of B-cell–predominant lymphoid infiltrates and lymphoepithelial lesions, lymphocytic mastitis and diabetic mastopathy do not appear to be associated with an increased risk for lymphoma.

Role of iron in T cell activation : TH1 clones differ from TH2 clones in their sensitivity to inhibition of DNA synthesis caused by IGG mabs against the transferrin receptor and the iron chelator deferoxamine

TH1 and TH2 helper T cell clones have been studied with respect to their sensitivity to inhibition of DNA synthesis by an IgG anti-transferrin receptor antibody (ATRA), the iron chelator deferoxamine, and the combination of the two reagents. TH1 clones are very sensitive to ATRA-mediated inhibition of DNA synthesis while TH2 clones are very resistant, but both TH1 and TH2 clones show significant down-modulation of surface transferrin receptors after ATRA exposure. TH2 clones exhibit larger chelatable iron storage pools than TH1 clones, however, and even partial chelation of TH2 cell storage iron does not fully convert a TH2 clone to the ATRA sensitivity pattern of a TH1 clone. It is therefore proposed that the greater resistance of TH2 clones to ATRA mediated inhibition derives from the combined effects of larger and less labile iron storage pools. These studies provide novel evidence indicating that nonuniform iron metabolism can exist within the T cell compartment and thus raise questions as to why such differences exist and how they can be integrated into models of the T cell activation process. These studies also suggest that the cell-mediated immune response in vivo, which is known to be sensitive to iron deficiency, may be evoked by effector cells which resemble TH1 clones insofar as iron metabolism is concerned.

Apparent movement due to closely spaced sequentially flashed dots in the human peripheral field of vision

Abstract Two dots in the peripheral field of vision, so close to one another that they are not resolved spatially when flashed together, induce a strong illusion of movement when they are flashed sequentially. Light adaptation changes the range of adequate interflash intervals. Stabilized vision, and a number of variants of the two-flash program, do not abolish the illusion whereas putative dichogeniculate presentation does. We have not been able to rule out the view that the illusion shares a common basis with the phenomena of metacontrast and real-movement perception.

Hepatocellular lymphoepithelioma-like carcinoma associated with epstein barr virus: a hitherto unrecognized entity.

ObjectiveLymphoepithelioma-like carcinoma (LELC) is an undifferentiated carcinoma with a dense lymphoid stroma. It has been reported in diverse organs and shows variable association with Epstein-Barr virus (EBV). Only a few EBV positive cases have been observed in the hepatobiliary system, all of which were considered to be cholangiocarcinomas. We report a unique case of hepatocellular LELC arising in a cirrhotic liver with EBV demonstrated in the tumor cells. Methods and ResultsA 39-year-old Hispanic female underwent an orthotopic liver transplant for end stage liver disease secondary to chronic hepatitis C. A high-grade hepatocellular carcinoma with a dense lymphocytic infiltrate was found in the explant as well as in a portal lymph node. Three months posttransplant, the patient developed numerous hepatic nodules with enlarged periaortic and portacaval lymph nodes. Biopsy of the hepatic nodules showed a recurrent hepatocellular carcinoma devoid of a dense lymphocytic infiltrate. Both the primary and recurrent tumors were positive for EBV by molecular studies. The patient eventually expired from liver failure over a 6-week period. ConclusionThis case represents the first report of EBV-positive hepatocellular LELC. It is particularly interesting given the precipitous clinical outcome, which was possibly related to immunosuppresive therapy.

Effects of Anti-Transferrin Receptor Antibodies on the Growth of Neoplastic Cells

One approach to creating a state of iron deprivation in tumors is to expose them to monoclonal antibodies against the transferrin receptor (ATRAs). This paper reviews the recent history of studies with ATRAs. Both multivalent (IgM or IgA) and bivalent (IgG) ATRAs exhibit anti-tumor activity in vitro and in vivo, but IgG ATRAs appear to be most effective when used with an iron chelator such as deferoxamine or when used in pairs. Much more information is needed in order to understand: (1) how ATRAs work by themselves and in conjunction with chelators; (2) why ATRAs differ from one another in terms of their inhibitory potency; (3) whether ATRAs can be used successfully in conjunction with other anti-tumor agents, and (4) why tumors exhibit marked differences in their sensitivity to the effects of ATRAs. The toxicity of iron deprivation arising from ATRA treatment alone seems modest, but only further experimental work in vivo and in phase-1 clinical trials can determine whether the most recent observations can be converted into truly useful therapeutic tools.

Inhibition of lymphocyte activation with anti-transferrin receptor Mabs: A comparison of three reagents and further studies of their range of effects and mechanism of action

Prior work has suggested that Mabs against the transferrin receptor (ATRAs) may function as selective inhibitors of lymphocyte activation and that T cell activation protocols may be more sensitive to ATRA-mediated inhibition than B cell activation protocols. New side-by-side functional comparisons of three ATRAs are presented. When these studies are considered with our prior work they demonstrate unambiguously that although one particular IgG ATRA consistently fails to inhibit LPS responses and although IgM ATRAs may be slightly more effective inhibitors than IgG ATRAs, ATRAs as a class consistently appear to abolish the MLR at submicrogram concentrations, essentially eliminate cytotoxic cell generation at concentrations between 1 and 5 micrograms/ml, and produce no more than about 50% inhibition of LPS responses at concentrations as high as 25 micrograms/ml. Therefore, an even stronger case can now be made for the idea that lymphocyte subsets differ in their dependence on transferrin receptor function during activation. This, in turn, makes an even stronger case for the idea that lymphocyte subsets differ in fundamental aspects of the management of their iron economies. New studies also show that IgG ATRAs appear to function by causing down-modulation of surface expression of the transferrin receptor in normal lymphocytes in a manner similar to that previously shown for tumor cells. It is clear that a sophisticated model will ultimately be required to account for all of the data arising from studies with ATRAs, and a new attempt at a more detailed construct is presented.