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The diploid eastern United States perennials-Verbesina helianthoides Michx, V. heterophylla (Chapm.) A. Gray, V. aristata (Ell.) A. A. Heller, and V. chapmanii Coleman-were successfully crossed in every combination. The F1 hybrids were vigorous, and mean pollen stainability ranged from 29% to 89%. Mean F1 female fertility ranged from 0 0% to 20 7%. The F2 and backcross plants were also vigorous and relatively fertile. Meiosis was highly regular in the parent species and F1's, with approximately 98% of the chromosomes occurring as bivalents in each case. The species are probably allopatric; however, the southern boundary of the range of V. aristata reaches to within approximately 15 miles of the range of V. chapmanii. The eastern boundary of V. aristata abuts on the western boundary of V. heterophylla, but their ranges apparently do not overlap. Topographic and edaphic factors which possibly maintain these species allopatric are discussed. The possible effects of Pleistocene fluctuations of temperature and...

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Development of resting and high-frequency-evoked metabolic patterns in the central nucleus of the inferior colliculus (ICC) in rat was investigated using the 2-deoxyglucose (2-DG) method. The effects of early short-term acoustic deprivation were studied by unilateral blockade of the external auditory meatus on postnatal day 12. Quantitative and qualitative analyses were performed on 2-DG uptake in 5 regional sectors of ICC. Adult resting ICC uptake is robust and exhibits regional differentiation with the ventromedial region preferentially labelled. At 17 days of age, resting uptake is substantially lower and regionally more uniform than in the adult. This age-dependent change is likely a function of the immature stage of neuronal and cochlear development at 17 days. Unilateral 50 kHz stimulation in the adult results in bilaterally enhanced 2-DG incorporation in ventromedial ICC, with a discrete band of elevated uptake along the contralateral ventromedial border. 50 kHz stimulation evokes ventromedial banding in the contralateral, but not ipsilateral, ICC of undeprived 17-day-olds, though at a much reduced level relative to the adult. Following monaural conduction blockade from 12 to 17 days of age, stimulation through the reopened deprived ear elicits a band of uptake along the lateral ICC border, oblique to the normal tonotopic gradient. Cochlear mechanisms may account for modified patterning of high-frequency-evoked 2-DG uptake in ICC subsequent to conductive hearing loss during a period of elevated susceptibility to experiential factors.

In a clinical trial based on the direct relationship between heart rate activity and energy expenditure during awake and asleep states, 21 healthy 18-month-old infants had their heart rate monitored over a period of 24 h and correlated with their nutritional intake, growth parameters, and measurements of adiposity (fat cell size and skinfold thickness). Infants with higher 24-h and asleep heart rates had smaller fat cells (r = -0.50) and skinfold thickness (abdominal r = -0.70, suprailiac r = -0.80, triceps r = -0.67). Infants with higher relative weight had higher heart rates during awake periods (r = 0.60). Infants sleeping for longer periods had higher weight gain velocity from birth (r = 0.51) and higher relative weight (r = 0.54). This study demonstrates the heart rate activity and duration of sleep are related to growth and adipose tissue deposition during infancy.

The course of development of the auditory brainstem response (ABR) in rat was examined. Pure tone pip stimuli of 3, 8 and 40 kHz at intensities up to 106 dB sound pressure level peak equivalent were presented to anesthetized rats on postnatal days (PND) 9, 10, 12, 14, 16, 20, 24, 36, 70 and 96. The peak-to-peak amplitude and the order of appearance of each wave was examined at each stimulus parameter. The first response to presentation of an auditory stimulus was observed on PND 12. This response was demonstrated to be of non-neural origin and presumed to be a summating potential representing hair cell function in the cochlea. The first neural responses, observed on PND 14, to 3- and 8-kHz tones only, appeared as two or more waves. High-frequency (40 kHz) tones did not elicit any substantive neural response on PND 14, but neural activity to this stimulus was observed consistently by PND 16. By PND 20, 4 waves were present consistently at all test frequencies, and by PND 36 the full adult complement of 5 vertex positive waves was recorded. The frequency-dependent sequential appearance of waves during development lends support to data charting the development of frequency mapping in the cochlea. The amplitude of waves I and II at 8 kHz decreased as the animal matured, while waves III-V generally showed an initial increase in amplitude at all test frequencies through PND 24 followed by a gradual amplitude decrease to adult levels. Possible sources for developmental changes in amplitude are discussed.

The indium trichloride method of Watson and Aldridge (38) for staining nucleic acids for electron microscopy was employed to study the relationship of DNA to the structure of the synaptinemal complex in meiotic prophase chromosomes of the domestic rooster. The selectivity of the method was demonstrated in untreated and DNase-digested testis material by comparing the distribution of indium staining in the electron microscope to Feulgen staining and ultraviolet absorption in thicker sections seen with the light microscope. Following staining by indium, DNA was found mainly in the microfibril component of the synaptinemal complex. When DNA was known to have been removed from aldehyde-fixed material by digestion with DNase, indium stainability was also lost. However, staining of the digested material with non-selective heavy metal techniques demonstrated the presence of material other than DNA in the microfibrils and showed that little alteration in appearance of the chromosome resulted from DNA removal. The two dense lateral axial elements of the synaptinemal complex, but not the central one to any extent, also contained DNA, together with non-DNA material.

A B S T R A C T Section Ximenesia is native to the southwestern United States and northern Mexico. Two of its taxa, Verbesina encelioides subsp. exauriculata and V. nana, are more western and occur in arid or semiarid regions, whereas the third taxon, V. encelioides subsp. encelioides, is more eastern and occurs in relatively moist regions. A crossing program revealed the absence of isolating barriers between the subspecies of V. encelioides. It is suggested that V. encelioides and V. nana are effectively isolated genetically by differences in ploidy. The study of several morphological characters of both field-collected and garden-grown populations indicates that differential adaptation for aridity has been of primary importance in the evolution of section Ximenesia. The taxa occupying arid regions show a reduction of transpiring surface, reduced reproductive capacity, and reduced dispersibility. IN THE MOST RECENT taxonomic treatment of section Ximenesia (Coleman, 1966a), two species were recognized. Verbesina encelioides (Cav.) Benth. & Hook., an annual, taprooted diploid (n = 17), was divided into two subspecies. Subspecies encelioides is eastern and lowland, occurring primarily in eastern Oklahoma, eastern Texas, and lowland northeastern Mexico. Subspecies exauriculata (Rob. & Greenm.) Coleman is western and highland and occurs throughout

A fraction of DNA, known to be replicating, was examined in the electron microscope. Characteristic branch points were identified as sites at which replication had been taking place. No localized strand separations, nor other structural alterations were discernible at these points.

Yeast display of antibody fragments has proven to be an efficient and productive means for directed evolution of single chain Fv antibodies for increased affinity and thermal stability, and more recently for the display and screening of a non-immune library. In this paper, we describe an elegant and simple method for constructing large combinatorial Fab libraries for display on the surface of Saccharomyces cerevisiae, from modestly sized, and easily constructed, heavy and light chain libraries. To this end, we have constructed a set of yeast strains and a two vector system for heavy chain and light chain surface display of Fab fragments with free native amino termini. Through yeast mating of the haploid libraries, a very large heterodimeric immune Fab library was displayed on the diploids and high affinity antigen specific Fabs were isolated from the library.

Binaural interactions were recorded in auditory cortical (AI) neurons of anesthetized adult cats that had unilateral atresias created shortly after birth by surgically ligating and cutting one external ear canal. At the time of the recording experiment, the atresia and associated debris were removed and tones were delivered to both tympanic membranes via a sealed and calibrated acoustic system. The majority of neurons recorded were in the cortex ipsilateral to the previously occluded ear. Thresholds for monaural stimulation of either the operated or unoperated ear were within normal range although thresholds to stimulation of the previously operated ear tended to occupy the upper end of the normal distribution. Monotonie and nonmonotonic spike count-vs-intensity functions derived from responses to monaural stimulation of the atretic ear were indistinguishable in their shape from those recorded in normal cats. All binaural classes were represented in our sample in proportions similar to those reported in cats with two normal ears. The forms of the functions relating spike count to interaural intensity differences and interaural phase differences were essentially the same as those seen in normal animals. The main binaural deficit observed under these conditions was a shift in the interaural intensity difference to which an AI neuron was most sensitive. The intensity needed for a stimulus to the atretic ear to participate in the binaural response was as much as 50 dB higher than that at the opposite normal ear in contrast to the nearly equal SPLs required for binaural interactions in cats with two ears intact. It was suggested that elevated thresholds at the previously operated ear could account for much of the shift observed.

The mean fat cell diameter was determined from measurements of abdominal adipose cells, obtained during inguinal hernia repair, of 126 white and 95 black males ranging in age from 1 through 48 months of age. The mean diameters of black and white subjects did not differ significantly, suggesting that differences in fatness among adults of these two ethnic groups have their origin beyond the age range of this study. The mean fat cell diameter increased through the 6-8 month age group, decreased until the end of the first year, and then levelled off through 48 months of age. Comparison of this curve with those for the triceps, subscapular, abdominal, and suprailiac skinfolds of the same subjects showed generally parallel courses except for the triceps, which continued to increase in size after the means of fat cell diameters and the other skinfolds had levelled off. Our data indicate that changes in body fatness on the trunk at least in the first 4 years of life may be accounted for by changes in fat cell size.

Patterns of normal development of the rat dorsal and ventral cochlear nuclei (DCN, VCN) and the changes induced in the adult by postnatal acoustic deprivation were examined using volumetric determinations. VCN and DCN showed striking increases in size during the interval from postnatal days 10 to 16 which includes meatus opening. Sound deprivation from postnatal day 10 resulted in the most pronounced reduction in both DCN and VCN. The findings suggest a sensitive period during which structural development in the cochlear nuclei is influenced by acoustic stimuli.

Tonotopic organization is a major feature of central auditory structures. The tritiated 2-deoxyglucose metabolic marking technique is demonstrated to be useful for visualizing isofrequency contours in the rat inferior colliculus. Following pure-tone stimulation, the central nucleus of the inferior colliculus shows discrete, prominent banding patterns and a well-organized tonotopic arrangement.

The effects of monaural deprivation by ligation of the external auditory meatus on spherical cells in the anteroventral cochlear nucleus of the rat were examined. Sound deprivation from postnatal days 10, 16, 24, and 36 resulted in a reduction in spherical cell size, with the most dramatic reduction produced by deprivation at 10 and 16 days after birth. The anteroventral cochlear nucleus was subdivided into dorsal, middle, and ventral zones which generally correspond to different frequency representations, and the effects of deprivation on cells in these regions were examined. Cells in the dorsal zone of the anteroventral cochlear nucleus were significantly smaller than cells in the middle or ventral zones, thus indicating a position-dependent effect of acoustic deprivation. Our results suggest a sensitive period during which alterations in normal acoustic input can significantly and differentially affect the development of cells in the anteroventral cochlear nucleus.

A technique is described which permits unambiguous identification of titanium-containing particles in thin sections of lung, with the use of a conventional transmission electron microscope and a conventional electron probe x-ray microanalyzer. Any titanium particle can be identified as titanium regardless of morphological changes. Chemical changes can be readily discovered by analysis of the x-ray spectrum that is emitted from the particle. The procedure is easily quantitated. The ability to examine thin sections by conventional transmission electron microscopy permits the identification of the cell sites in which titanium is located with sufficient resolution to determine whether cell origanelles or cytoplasmic membranes are involved. (JTE)

Autoradiography was used to assess the incorporation of [2-14C]deoxy-D-glucose by the auditory brain stem of young and aging mice of the C57BL/6 strain (which demonstrates progressive chronic sensorineural hearing loss) and the CBA strain (which maintains good hearing until late in life). Animals were injected with labeled 2-deoxyglucose and placed in quiet for 45 min; brain stem sections were prepared for autoradiography. The amounts of 2-deoxyglucose incorporated into the anterior ventral cochlear nucleus (AVCN), inferior colliculus (IC), and trigeminal nerve (TN) were densitometrically analyzed. Within each subject, the densities of the three structures were statistically compared. In every mouse, inferior colliculus density was greater than that of the anterior ventral cochlear nucleus, which was greater than trigeminal nerve density. To compare subject groups, relative densities (inferior colliculus and anterior ventral cochlear nucleus re: trigeminal nerve) were used; no significant differences were found between groups. Thus, aging, with or without severe loss of hearing, is not associated with altered incorporation of 2-deoxyglucose (and presumably glucose) in quiet.

The most common source of uncertainty in the use of electron beam analytical techniques with biological materials is the preparation technique: the steps that intervene between the normal in vivo condition of the sample and the state in which it is analyzed. Generally, these steps involve removing material from an aqueous, atmospheric pressure environment to one that is dry and at relatively high vacuum, and may also include sectioning, sawing, or fracturing in order to expose the portions of interest. During this treatment, material must not be lost, redistributed, or gained, and the original structural relationships of the material must be conserved. These are quite restrictive requirements, and are usually satisfied only in part. Indeed, establishing the degree to which they are met is often a major problem in itself. There may be no universally satisfactory preparation technique, although some of the freezing techniques approach this. Thus it is usually the responsibility of each analyst to establish criteria for the problem of interest. In general, the properties of the sample determine which preparative techniques can be employed. Thus, if one wishes to analyze tissue fluids, crystalline inclusions, or cell organelles, it is likely that different methods will be useful. A brief survey of some of the methods that are available follows, along with some suggestions about their advantages as well as the hazards that are likely to be associated with them. A general overview was given by Coleman and Terepka.(1)

The electrical activity of the dorsal and ventral hippocampus and of the anterior and posterior neocortex is described and illustrated during the development of trained operant behavior in water-deprived cats learning to bar-press for water reward under three operant conditioning schedules: (a) continuous reinforcement; (b) alternating 10 sec periods of reinforcement and non-reinforcement; and (c) mixed reinforcement and non-reinforcement with special auditory and visual cues. In addition, the contrasting effects of 100 c/sec electrical stimulation of the medial and lateral hypothalamic systems upon hippocampal electrical activity and operantly trained behavior are reported. During the early stages of learning to bar-press for water reward, when close attention to the bar and water-well are required, hippocampal electrical activity manifests synchrony (theta rhythm) and neocortical electrical activity is desynchronized. Subsequently, as bar-pressing performance improves and requires little attention to the manipulanda, the pattern of hippocampal and neocortical electrical activity is one of irregular slow waves mixed with low voltage high frequency activity, characteristic of relative inattention and automatic performance. During alternating or mixed reinforcement and non-reinforcement schedules of operant training differential effects are observed during reinforcement and non-reinforcement periods, the latter being characterized generally by lower voltage, mixed low and high frequency activity, except when orienting and shifting of attention occurred with associated theta rhythm bursts. Stimulation of the medial hypothalamic system has a striking inhibitory effect upon bar-pressing for water reward. Bar-pressing ceases for many minutes but its eventually resumed at the pre-stimulation rate. In contrast, stimulation of the lateral hypothalamic system only interrupts bar-pressing for a matter of seconds. Possible causes of these differential effects, and especially the prolonged inhibition of bar-pressing induced by medial hypothalamic stimulation, are discussed.

Brainstem and forebrain projections to major subdivisions of the rat inferior colliculus were studied by using retrograde and anterograde transport of horseradish peroxidase. Retrograde label from injection into the external cortex of the inferior colliculus appears bilaterally in cells of the inferior colliculus, as well as in other brainstem auditory groups including the ipsilateral dorsal nucleus of the lateral lemniscus and contralateral dorsal cochlear nucleus. The external cortex is the only collicular subdivision where an injection labels cells in the contralateral cuneate nucleus, gracile nucleus, and spinal trigeminal nucleus. Other projecting cells to the external cortex are found in the lateral nucleus of substantia nigra, the parabrachial region, the deep superior colliculus, the midbrain central gray, the periventricular nucleus, and area 39 of auditory cortex. Injection of the dorsal cortex of inferior colliculus heavily labels pyramidal cells of areas 41, 20, and 36 of the ipsilateral neocortex. Anterograde label from a large injection of auditory cortex is densely distributed in the dorsal cortex, lesser so in the external cortex, and only slightly in the central nucleus. Labelled cells appear in the central nucleus, dorsal cortex, and external cortex, primarily ipsilaterally, following dorsal cortex injection. Relatively few cells from other brainstem auditory groups show projections to the dorsal cortex. Injection of the central nucleus of the inferior colliculus results in robust labelling of nuclei of the ascending auditory pathway including the anteroventral, posteroventral, and dorsal cochlear nuclei (mainly contralaterally), and bilaterally the lateral superior olive, lateral nucleus of the trapezoid body, dorsal nucleus of the lateral lemniscus, and the central nucleus, dorsal cortex, and external cortex of the colliculus. The medial superior olive, superior paraolivary nucleus, and ventral nucleus of the trapezoid body essentially show ipsilateral projections to the central nucleus. The differential distribution of afferents to the inferior colliculus provides a substrate for functional parcellation of collicular subdivisions.

Thirty-three male subjects participated in a study to examine the effect of supplements of multiple vitamins and minerals, local therapy (periodontal instrumentation and oral hygiene instruction) and a combination of both on gingival inflammation and bacterial plaque formation. Subjects were given either multivitamin and mineral supplements or placebos on a double-blind basis for 21 days. On Day 7, the mandibular incisors were instrumented, and each subject was instructed in brushing and flossing. Observations were taken at Days 0, 7 and 21. There was a significant (P = 0.004) effect from micronutrient supplementation at Day 7 on the gingival index but no significant effect on the plaque index. On Day 21 there was no statistical superiority noted for the supplemented group in respect to either the gingival or plaque index, although the gingival index approached significance (P = 0.062).

The sizes of large spherical and fusiform cells of the anteroventral cochlear nucleus were studied in rats reared after unilateral middle ear ossicle removal on postnatal days 10 or 16, or after bilateral removal on postnatal day 16. In animals deprived monaurally from 10 days after birth, large spherical cell areas of the deprived side were reduced by 21% compared to the nondeprived side; the deprived cells were 17% smaller than cells of control animals. Large spherical cells contralateral to the deprived ear were slightly larger than those of controls. Large spherical cells monaurally deprived from the 16th day showed effects similar to those deprived from the onset of hearing, although cell reduction on the deprived side was only 10% relative to controls. In contrast to the consequences of monaural sound deprivation, binaural deprivation from the 16th day resulted in no reduction in size of large spherical cells. Fusiform cells of animals in monaurally and binaurally deprived groups were not significantly smaller than the fusiform cells of controls. Ear histologies confirmed that each cochlea and spiral ganglion was normal in experimental and control animals. Because large spherical cells receive unilateral input directly from the acoustic nerve and are known to project bilaterally to the medial superior olive, the bilateral differences observed in large spherical cell area following rearing under monaural deprivation conditions may reflect a form of imbalanced binaural interactions between axon terminals of spherical cells.

An attempt is made to correlate structure and transport function in the embryonic chorioallantoic membrane. The fine structure of the endoderm and ectoderm in the membrane was examined with particular attention given to the morphological changes that occur when transport is established,in vivo. Two distinctive cells, a granule-rich cell and a mitochondria-rich cell, appear in the endoderm at the time allantoic fluid sodium, chloride and water reabsorption commences. These are indistinguishable from the cells described in toad bladder epithelium. It is suggested that the granule-rich cell is responsible for bulk water movement and the mitochondria-rich cell is specifically engaged in active sodium transport. In the ectoderm, two distinctive cell types are also found to be associated with the onset of active calcium transport. These are referred to as the capillary-covering cell and the villus-cavity cell. The preponderate capillary-covering cell is most likely responsible for transcellular calcium transport. It is postulated that the function of the villus-cavity cell is to secrete hydrogen ions which are necessary, along with carbonic anhydrase, to mobilize Ca(++) from the insoluble calcium carbonate of the eggshell.

Procedures were developed for preparation of red cell membranes almost free of hemoglobin but with minimal loss of membrane proteins. Two water-soluble protein fractions are described, each constituting about 25% of the ghost protein. The first is ionically bonded and can be solubilized in water rapidly at pH 7.0 and more slowly at higher ionic strength solutions, with a minimal rate at 20MM. This fraction contains four major components with molecular weights ranging from 30,000 to 48,000. The second fraction can only be solubilized at an appreciable rate if Ca(++) is absent and at higher pH (9.0). It is predominantly a single molecular weight component (150,000). It tends to aggregate at higher ionic strength and in the presence of Ca(++). Evidence is presented suggesting that the water-soluble proteins are present at the inner face of the membrane. The lipids remain in a water-insoluble residue that contains four major protein components ranging in molecular weight from 30,000 to 100,000. The latter is the predominant component. Only the residue contains the Na(+)-K(+)-activated ATPase, the cholinesterase, antigenic activity and most of the sialic acid and carbohydrate. The first water-soluble fraction contains a Mg(++)-activated ATPase. The extraction of the water-soluble proteins is accompanied by anatomical changes resulting finally in the formation of small membranous vesicles.

The chorioallantoic membrane of the developing chick embryo is an epithelium that actively transports calcium. The methodology utilized to prepare this soft tissue for calcium localization with the electron probe x-ray microanalyzer is presented in detail. The preparative procedures are evaluated according to general histochemical principles and in relationship specifically to electron probe investigations. It is shown that the method employed in these studies preserves the normal fine structure of the tissue, prevents selective loss of calcium, permits only minor losses of total calcium and appears to maintain the distribution of calcium that existed in vivo. Examples are presented of artifacts that can be induced during tissue sectioning and mounting procedures. Problems of defining electron probe resolution in biologic specimens are discussed, and the critical importance of evaluating x-ray images in association with simultaneously generated sample current images is emphasized.

This research examined the impact of in-vehicle information system (IVIS) interactions on the driver’s cognitive workload; 257 subjects participated in a weeklong evaluation of the IVIS interaction in one of ten different model-year 2015 automobiles. After an initial assessment of the cognitive workload associated with using the IVIS, participants took the vehicle home for 5 days and practiced using the system. At the end of the 5 days of practice, participants returned and the workload of these IVIS interactions was reassessed. The cognitive workload was found to be moderate to high, averaging 3.34 on a 5-point scale and ranged from 2.37 to 4.57. The workload was associated with the intuitiveness and complexity of the system and the time it took participants to complete the interaction. The workload experienced by older drivers was significantly greater than that experienced by younger drivers performing the same operations. Practice did not eliminate the interference from IVIS interactions. In fact, IVIS interactions that were difficult on the first day were still relatively difficult to perform after a week of practice. Finally, there were long-lasting residual costs after the IVIS interactions had terminated. The higher levels of workload should serve as a caution that these voice-based interactions can be cognitively demanding and ought not to be used indiscriminately while operating a motor vehicle.

Innovations: Self-instructed use of microcomputers in the library

Section Ximenesia is treated as composed of two species, Verbesina encelioides (Cav.) Benth. & Hook. f. and V. nana (Gray) Rob. & Greenm. The annual V. encelioides is native to the southwestern U. S. and the Mexican Plateau, but is weedy and has extended its range into several regions of the world. It is composed of an eastern, typical subspecies and a western subspecies, exauriculata (Rob. & Greenm.) Coleman, stat. nov. In this species n 1 17. V. nana is perennial, is mostly restricted to the Trans-Pecos region of Texas and New Mexico and is n 34. Section Ximenesia is a small group native to the southwestern United States and the Mexican Plateau, the species of which occur primarily in arid grass- and scrublands. One species, Verbesina encelioides (Cav.) Benth. & Hook., is weed,y and has extended its range into several regions of the world. This same species is cultivated as an ornamental, and is of further economic importance because of its poisonous, even lethal, properties to cattle and sheep (White, 1938). Robinson and Greenman (1899), in their comprehensive treat- ment of Verbesina, recognized three species in section Ximenesia. One of these, V. encelioides, was treated as composed of three varie- ties. In the present treatment the species are reduced to two and V. encelioides is considered as composed of two subspecies.