Recurrent isolation of an uncommon yeast, Candida pararugosa, from a sarcoma patient.

A yeast was repeatedly isolated from the saliva of a sarcoma patient. A relatively uncommon species, Candida maris, was identified based on the API 20C profile. The yeast species most frequently obtained from the patient's mother and from clinic staff was Candida albicans. A comparison of the yeast obtained from the patient with the type strain of C. maris strongly suggested that the former was not representative of C. maris. Analysis of partial ribosomal DNA sequences of the patient strain and from the type strain of C. maris showed that the two are phylogenetically not closely related. The patient strain was very close to Candida pararugosa, a relatively uncommon asporogenous yeast. DNA reassociation studies among C. pararugosa and patient isolates showed that they were conspecific. We could not determine the source of the yeast infection. This case will alert hospital staff to be aware of the possibility of unexpected environmental microorganisms as causes of infections, colonizations and persistent environmental contamination events in immunocompromised patients.

A limited repertoire of mutations of the luteinizing hormone (LH) receptor gene in familial and sporadic patients with male LH-independent precocious puberty.

Herein, we report mutation analysis of the LH receptor gene in 17 males with LH-independent precocious puberty, of which 8 were familial and 9 had a negative family history. A total of 7 different mutations (all previously reported) were detected in 12 patients. Among 10 European familial male-limited precocious puberty (FMPP) patients who had a LH receptor gene mutation, none had the Asp578Gly mutation, which is responsible for the vast majority of cases in the U.S. The restricted number of activating mutations of the LH receptor observed in this and other studies of FMPP strongly suggests that an activating phenotype is associated with very specific sites in the receptor protein. Clinical follow-up of the 5 patients who did not have LH receptor mutations shows that such cases most likely do not have true FMPP. LH receptor mutation analysis provides a sensitive tool for distinguishing true FMPP from other causes of early-onset LH-independent puberty in males.

A missense mutation in the second transmembrane segment of the luteinizing hormone receptor causes familial male-limited precocious puberty.

Patients with familial male-limited precocious puberty present with early onset of puberty. Several missense mutations in the LH receptor gene that cause amino acid substitutions in the sixth transmembrane segment of the receptor protein have been shown to be a cause of the disorder. We have identified a novel LH receptor gene mutation in a patient with familial male-limited precocious puberty that results in a threonine for methionine substitution at position 398 in the second transmembrane segment of the receptor protein. In vitro expression in human embryonic kidney 293 cells of this LH receptor mutant and two previously described LH receptor mutants showed that cAMP production in the absence of hormone was elevated up to 25-fold compared to the basal level of the wild-type receptor. The ED50 values of hormone-induced cAMP production were within the same range for wild-type and mutant receptors, but maximal hormone-induced cAMP production was relatively low for mutant receptors. We also produced receptors containing amino acid substitutions in both the second and sixth transmembrane segments. For these double mutants, basal receptor activities were similar to the basal activities observed in single mutants, whereas hormone-induced receptor activation was almost completely abolished.

Selectivity for temporal characteristics of sound and interaural time difference of auditory midbrain neurons in the grassfrog: a system theoretical approach.

The selectivity for temporal characteristics of sound and interaural time difference (ITD) was investigated in the torus semicircularis (TS) of the grassfrog. Stimuli were delivered by means of a closed sound system and consisted of binaurally presented Poisson distributed condensation clicks, and pseudo-random (RAN) or equidistant (EQU) click trains of which ITD was varied. With RAN and EQU trains, 86% of the TS units demonstrated a clear selectivity for ITD. Most commonly, these units had monotonically increasing ITD-rate functions. In general, units responding to Poisson clicks, responded also to RAN and EQU trains. One category of units which showed strong time-locking had comparable selectivities for ITD with both stimulus ensembles. A second category of units showed a combined selectivity for temporal structure and ITD. These units responded exclusively to EQU trains in a nonsynchronized way. From the responses obtained with the Poisson click ensemble so-called Poisson system kernels were determined, in analogy to the Wiener-Volterra functional expansion for nonlinear systems. The kernel analysis was performed up to second order. Contralateral (CL) first order kernels usually had positive or combinations of positive and negative regions, indicating that the contralateral ear exerted an excitatory or combined excitatory-inhibitory influence upon the neural response. Ipsilateral (IL), units were characterized by first order kernels which were not significantly different from zero, or kernels in which a single negative region was present. A large variety of CL second order kernels has been observed whereas rarely IL second order kernels were encountered. About 35% of the units possessed nonzero second order cross kernels, which indicates that CL and IL neural processes are interacting in a nonlinear way. Units demonstrating a pronounced selectivity for ITD, were generally characterized by positive CL combined with negative IL first order kernels. Findings suggested that, in the grassfrog, neural selectivity for ITD mainly is established by linear interaction of excitatory and inhibitory processes originating from the CL and IL ear, respectively. Units exhibiting strong time-locking to Poisson clicks and RAN and EQU trains had significantly shorter response latencies than moderately time-locking units. In the first category of units, a substantial higher number of nonzero first and second order kernels was observed. It was concluded that nonlinear response properties, as observed in TS units, most likely have to be ascribed to nonlinear characteristics of neural components located in the auditory nervous system peripheral to the torus semicircularis.

Sensitivity for interaural time and intensity difference of auditory midbrain neurons in the grassfrog.

The sensitivity for interaural time (ITD) and intensity (IID) difference was investigated for single units in the auditory midbrain of the grassfrog. A temporally structured stimulus was used which was presented by means of a closed sound system. At best frequency (BF) the majority of units was selective for ITD as indicated by an asymmetrically (73%) or symmetrically (7%) shaped ITD-rate histogram. About 20% appeared to be nonselective. Units with a symmetrical rate histogram had BFs well above 0.9 kHz, whereas for the other categories no relationship with BF was observed. Most units had a selectivity for ITD which was rather independent from frequency and absolute intensity level. In 62% of the units interaural time difference could be traded by interaural intensity difference. In most cases this so-called time-intensity trading could be explained by the intensity-latency characteristics of auditory nerve fibres. About 20% was sensitive to IID only and 5% to ITD only. A binaural model is proposed which is based on the intensity-rate and intensity-latency characteristics of auditory nerve fibres, the linear summation of excitatory and inhibitory post synaptic potentials in second order neurons, and spatiotemporal integration at the level of third order neurons. By variation of only a small number of parameters, namely strengths and time constants of the connectivities, the range of experimentally observed response patterns could be reproduced.

[Late results following endoprosthetic lunate bone replacement]. / Spätergebnisse nach endoprothetischen Lunatumersatz.

Long-term results of silastic lunate spacer implantation are reported especially concerning late complications in the form of reactive synovitis and intraosseous cysts. As result one case of silicon cyst and no silicon synovitis was found.

Influence of adaptation on neural sensitivity to temporal characteristics of sound in the dorsal medullary nucleus and torus semicircularis of the grassfrog.

The influence of long-term (approximately minutes) adaptation on the responses of 15 dorsal medullary nucleus (DMN) and 36 torus semicircularis (TS) neurons of the grassfrog to temporally structured stimuli has been investigated. The neurons were stimulated with periodic click trains and sine amplitude modulated (AM) tone bursts presented in isolation or embedded in a randomly structured background. A tone burst with the envelope of the conspecific mating call was presented in addition. The stimulus-response relation has been analyzed by peri-stimulus time histograms (PSTH), average rate and synchronization index histograms. Global response types based on average rate and synchronization index were altered by adaptation in about half of the units. The trend was to enhance selectivity for temporal frequency and to increase response synchronization. Long-term adaptation had a moderate to dramatic effect on detailed response structure, as expressed by the PSTH, in 86% of the DMN and 96% of the TS units. In general the adaptational influence was somewhat larger for AM tones than for clicks. The observed effects could not be explained by a simple neural threshold shift. In almost all units the response to periodic sound bursts can be regarded as a modulation below and above an average response level induced by the random background. The response to the mating call resemblant tone burst was clearly distinct neural characteristics on long-term adaptation, has important implications for system-theoretical stimulus-response analysis procedures.

A combined sensitivity for frequency and interaural intensity difference in neurons in the auditory midbrain of the grassfrog.

The relation between spectral tuning and sensitivity for interaural intensity difference (IID) was studied for single units in the auditory midbrain of the grassfrog. The stimuli consisted of sequences of pure tones of different frequency and interaural intensity differences presented by means of a closed sound system. At best excitatory frequency, three types of binaural interaction were observed: E0 (one ear excitatory 23%), EE (both ears excitatory 9%) and EI (one ear excitatory, the other inhibitory 67%). For a considerable number of units different types of binaural interaction were observed for different stimulus frequencies. More than 30% of the binaural units had multiple excitatory and inhibitory regions in their spectrotemporal selectivity. E0 and EI units had uniformly distributed best frequencies, EE units generally had best frequencies near 1.0 kHz. The E0 and EE categories had response latencies less than about 70 ms whereas EI units could have longer latencies. Most EE and all EI category units had sigmoidally shaped IID-rate curves. About 40% of the units had a combined sensitivity for sound spectrum and IID which was invariant to overall stimulus intensity. For nearly all EI units the inhibitory influence of the ipsilateral ear was confined to frequencies in the 0.4-1.6 kHz range and was not correlated with a unit's best frequency. By means of a simple additive model we demonstrated that determination of sound source laterality can be achieved by ensemble coding in the auditory midbrain.

Responses of auditory brainstem neurons in the grassfrog to clicks.

Single unit recordings were made in the dorsal medullary nucleus and in the torus semicircularis of the immobilized grassfrog. The natural calls have a periodic pulsatile structure. To investigate the coding of pulse repetition rate periodic click trains with varying pulse repetition rate and an ensemble of clicks distributed randomly in time were used as stimuli. In the dorsal medullary nucleus strong time-locking to clicks was found. Most units showed an activation followed by suppression response. Some units showed a preference for pulse repetition rates matching their low-frequency sensitivity. In the torus semicircularis part of the units showed responses similar to dorsal medullary nucleus units. Other response types were activation irrespective of pulse repetition rate, and suppression followed by activation. The responses to the two stimulus ensembles were more compatible in the dorsal medullary nucleus than in the torus semicircularis.

Coherent neural activity in the auditory midbrain of the grassfrog.

With a dual-electrode configuration separable few-unit activity was recorded both on one electrode as well as on two electrodes in the auditory midbrain of the grassfrog to a large variety of stimuli. Activity recorded on one electrode was separated by a pattern recognition technique through the use of features of the action potential waveform. Functional connections between units were established on basis of cross-correlation histograms of pairs of simultaneously recorded units. A hierarchical scheme was adopted to describe the various manifestations of neural correlation. If a peak or trough was observed in the simultaneous cross-correlation histogram, irrespective of stimulus conditions, this was called neural synchrony. If this peak or trough was not equal to its shift predictor estimating the stimulus contribution, neural correlation was considered to be present. About 60% of the pairs exhibited neural synchrony, mostly due to shared stimulus influences, independent of mutual distance of units. About 15% of the unit pairs showed neural correlation indicating a functional neural connection. Neural correlation was observed only in units with a distance smaller than 300 micron. The majority (approximately 85%) of the cases showing neural correlation could be ascribed to neural shared input. Unidirectional excitation was observed only in unit pairs recorded on the same electrode. Unidirectional inhibition could not be demonstrated. The dependency of occurrence of neural correlation on unit distance has implications for models of the functional organization of the auditory midbrain. About half of the neurally correlated pairs showed stimulus dependencies of their functional connections. Together with the observed lack of stimulus invariance of single-unit spectrotemporal sensitivities this indicates a dynamic stimulus dependency of functional neuronal organization in the auditory midbrain of the grassfrog. Neuron pairs with a large overlap of their spectrotemporal sensitivities on average had neurally correlated activities more often than pairs with a smaller amount of overlap. In comparison to single-unit coding, ensemble coding by populations of neurons may show an enhanced selectivity to stimulus characteristics.

Detection and estimation of neural connectivity based on crosscorrelation analysis.

Crosscorrelation analysis of simultaneously recorded activity of pairs of neurons is a common tool to infer functional neural connectivity. The adequacy of crosscorrelation procedures to detect and estimate neural connectivity has been investigated by means of computer simulations of small networks composed of fairly realistic modelneurons. If the mean interval of neural firings is much larger than the duration of postsynaptic potentials, which will be the case in many central brain areas excitatory connections are easier to detect than inhibitory ones. On the other hand, inhibitory connections are revealed better if the mean firing interval is much smaller than post-synaptic potential duration. In general the effects of external stimuli and the effects of neural connectivity do not add linearly. Furthermore, neurons may exhibit a certain degree of timelock to the stimulus. For these reasons the commonly applied "shift predictor" procedure to separate stimulus and neural effects appears to be of limited value. In case of parallel direct and indirect neural pathways between two neurons crosscorrelation analysis does not estimate the direct connection but instead an effective connectivity, which reflects the combined influences of the parallel pathways.

Coincidence detection in auditory neurons: a possible mechanism to enhance stimulus specificity in the grassfrog.

It is still a matter of debate whether neurons in the higher central nervous system of anurans become progressively more sharply tuned to sounds that have a behavioral importance or that such coding is performed by (small) groups of neurons. The approach we have taken to investigate this matter comprises simultaneous single-unit recording using two microelectrodes in the auditory midbrain of the grassfrog. The present study deals with 96 pairs of units responding to an ensemble of natural and synthetic mating calls in which carrier frequency and pulse-repetition rate were varied. This ensemble was presented without noise and also with background noise of increasing intensity. The spike trains were analysed for correlations between their firings. In 34 pairs (35%) a functional connection, mostly common input, was present. By selecting one of the units of a pair as a trigger it was investigated which window for a coincidence analysis would result in enhanced specificity for the unit pair. Such an analysis based on a logical AND operation could be a model for the action of a neuron on which both units under study would converge, and which would then show an enhanced specificity in their response to a stimulus ensemble. It was found that in 20 pairs (21%) the logical AND operation was more selective than each of the component neurons. The largest time window for which the selectivity was found was evenly distributed over the values 8 ms, 32 ms and 128 ms, in one case selectivity was found only for a window of 2 ms. There was neither preference for selective pairs to be found for recordings with one electrode (45 cases) or dual electrodes (51), nor for independent (62) versus functionally connected (34) pairs. In some cases selectivity resulted in a preference for one specific call, in other cases it resulted in a loss of responsiveness to the masking noise effectively resulting in an enhanced signal-to-noise ratio. The analysis stresses the importance of spatiotemporal patterns of nervous activity for the representation of sounds in the auditory midbrain of anurans.

Sensitivity of neurons in the auditory midbrain of the grassfrog to temporal characteristics of sound. I. Stimulation with acoustic clicks.

The coding of fine-temporal structure of sound, especially pulse repetition rate, was investigated on the single-unit level in the auditory midbrain of the grassfrog. As stimuli periodic click trains and Poisson distributed click ensembles have been used. The response to periodic click trains was studied in two aspects, focussing on two types of possible codes: a rate code and a synchrony code. From the iso-intensity rate histogram five basic average response rate characteristics as function of pulse repetition rate have been established: low-pass, band-pass, high-pass, bimodal and non-selective unit types. The synchronization capability, expressed in a synchronization index, was for a small majority of units non-significant and a low-pass function of pulse repetition rate for most of the other units. The rate code showed the largest diversity of response types and an enhanced selectivity to pulse repetition rate. The stimulus-response relation to Poisson distributed click ensembles was investigated by a non-linear system theoretical approach. On the basis of first- and second-order Poisson kernels possible neural mechanisms accounting for temporal selectivity were determined. A considerable fraction of units exhibited response characteristics that were invariant to changes in sound pressure level and average click rate. These units may function as feature detectors of fine-temporal structure of sound. The spectro-temporal sensitivity range of the auditory midbrain of the grassfrog appeared to be broad and not particularly tuned to the ensemble of conspecific cells.

Sensitivity of neurons in the auditory midbrain of the grassfrog to temporal characteristics of sound. II. Stimulation with amplitude modulated sound.

The coding of fine-temporal structure of sound, especially of frequency of amplitude modulation, was investigated on the single-unit level in the auditory midbrain of the grassfrog. As stimuli sinusoidally amplitude modulated sound bursts and continuous sound with low-pass Gaussian noise amplitude modulation have been used. Both tonal and wideband noise carriers have been applied. The response to sinusoidally amplitude modulated sound bursts was studied in two aspects focussing on two types of possible codes: a rate code and a synchrony code. From the iso-intensity rate histogram five basic average response characteristics as function of modulation frequency have been observed: low-pass, band-pass, high-pass, bimodal and non-selective types. The synchronization capability, expressed in a synchronization index, was non-significant for 38% of the units and a low-pass function of modulation frequency for most of the other units. The stimulus-response relation to noise amplitude modulated sound was investigated by a non-linear system theoretical approach. On the basis of first- and second-order Wiener-Volterra kernels possible neural mechanisms accounting for temporal selectivity were obtained. About one quarter of the units had response characteristics that were invariant to changes in sound pressure level and spectral content of the carrier. These units may function as feature detectors of fine-temporal structure of sound. The spectro-temporal sensitivity range of the auditory midbrain of the grassfrog appeared not to be restricted to and showed no preference for the spectro-temporal characteristics of the ensemble of conspecific calls. Comparison of response characteristics to periodic click trains as studied in the companion paper (Epping and Eggermont, 1986) and sinusoidally amplitude modulated sound bursts revealed that the observed temporal sensitivity is due to a combination of sensitivities to sound periodicity and pulse duration. It was found that for most units the first-order kernels for Gaussian amplitude modulated stimuli and Poisson distributed click stimuli were alike. In contrast second-order kernels for the Gaussian amplitude modulated stimuli often represented only static non-linearities, while second-order kernels for Poisson distributed clicks (Epping and Eggermont, 1986) mostly revealed dynamic non-linearities.

Sensitivity of neurons in the auditory midbrain of the grassfrog to temporal characteristics of sound. III. Stimulation with natural and synthetic mating calls.

Using an ensemble of natural and synthetic mating calls we studied the single unit (N = 189) and small neuronal group (91 unit pairs) responsiveness in the auditory midbrain of the grassfrog. We compared this responsiveness to that obtained for toneburst and sine-modulated stimuli in order to reveal the presence of so-called mating call detectors. Under the set of stimuli used 5% of the single units appeared to respond selectively to the natural mating call. Simultaneously recorded units appeared to have in about half of the cases identical properties, in the other half even completely complementary properties were found. These properties are related to the organizational structure of the torus semicircularis. A mechanism of feature extraction based on near coincident firings in simultaneously recorded neurons is presented.

Single-unit characteristics in the auditory midbrain of the immobilized grassfrog.

The anuran auditory midbrain of the grassfrog (Rana temporaria L.) was studied by a combined spectro-temporal analysis of sound preceding neural events. From the spectro-temporal sensitivities (STS) estimates of best frequencies (BF) and latencies (LT) were derived. Several types of STSs were observed: monomodal excitatory STSs comprised about half of the cases. Bimodal excitatory STSs, i.e. STSs with two discrete excitation regions, were observed in about 25%. Trimodal and broadly tuned STSs comprised about 5%. The remaining 20% of the STSs were characterized by inhibitory phenomena such as pure inhibition, sideband inhibition and post-activation inhibition. The distribution of best frequencies matches the frequency spectrum of the animal's vocalizations. A relative absence of monomodal units was noted in the mid frequency range. The distribution of latencies was bimodal over the range 7-108 ms. For each unit 6 functional parameters were determined; besides BF and LT these were: form of the STS (i.e. monomodality versus multimodality), spontaneous activity, binaural interaction, and firing mode (i.e. sustained versus transient) upon continuous noises stimulation. In addition, two structural parameters were considered: location in the torus and action potential waveform. Large correlations appeared between LT and action potential waveform, and between BF and binaural interaction type. Tonotopy was not found. A comparison was made between results from this study with a previous study on lightly anesthetized grassfrogs, using the same stimulus paradigms (D.J. Hermes et al. (1981): Hearing Res. 5, 147-178; D.J. Hermes et al. (1982): Hearing Res. 6, 103-126). Spontaneous activity, inhibitory phenomena and complex STSs were common using immobilization, whereas these have hardly been observed using anesthesia. Furthermore, interdependencies between the neural characteristics are substantially weaker for the immobilized preparation.

Relation of binaural interaction and spectro-temporal characteristics in the auditory midbrain of the grassfrog.

The relation between binaural interaction type and spectro-temporal characteristics was studied for single units in the auditory midbrain of the grassfrog. Tonal and continuous wideband noise ensembles have been used as stimuli. Spectro-temporal sensitivities were determined for ipsi-, contra- and bilateral stimulus presentation by a closed sound system. Binaural interaction was classified in monaural EO (one ear excitatory), binaural EE (both ears excitatory) and EI (one ear excitatory, the other inhibitory) and purely inhibitory categories. Binaural interaction appeared to be rather invariant to alterations in stimulus intensity and type. A very clear correlation was observed between best frequency and binaural interaction type: EE units are predominantly of high best frequency, whereas EI units are predominantly of low best frequency. The correlation with latency was less significant: EE units tended to have somewhat shorter latencies that EI units. EO units take an intermediate position. Comparisons of ipsi-, contra- and bilateral spectro-temporal sensitivities, revealed differences in best frequency, latency and temporal discharge pattern. In some units a complex interplay of excitatory and inhibitory monaural influences was demonstrated. A number of units was recorded, which were characterized by multiple activation or suppression areas. The majority of these units exhibited frequency-dependent binaural interaction types. In some units it was noticed that binaural interaction type can be dependent on state of adaptation. A comparison of binaural interaction types of neighbouring units provided only weak evidence for a binaural organization in the anuran auditory midbrain, since simultaneously recorded pairs shared the same binaural interaction type only slightly more than expected by mere chance (chi 2-test, P less than 0.10).

Directional hearing in the grass frog (Rana temporaria L.): I. Mechanical vibrations of tympanic membrane.

The vibration characteristics (amplitude and phase as a function of frequency) of the tympanic membrane in the grass frog were measured using a laser-doppler velocity meter. It was tested to what extent the frog's acoustic system behaves as a pressure gradient receiver. This might clarify how the frog localizes sound. Using a closed sound system the membrane was stimulated at three different entrances: in front of the membrane, at the contralateral ear and from inside the mouth. A combination of these can describe the motion of the membrane under free field conditions. It is found that the sound entrance from inside the mouth will give almost identical vibration characteristics as stimulation in front of the membrane. This can yield a perfect gradient receiver mechanism, when the frog opens its mouth. It is doubted however whether the frog in nature needs to open its mouth for localization of sound. With mouth closed the effectiveness of the gradient receiver will be determined by the transmission characteristics of sound across the tissues of the mouth. The entrance of sound via the contralateral ear is only effective at frequencies between 800 and 1600 Hz. At those frequencies crosstalk between the membranes is however not more than -4 to -8 dB. This is subject to changes in the acoustic properties of the mouth cavity and can possibly be altered by the frog in free nature.