Molecular bases of odor discrimination: Reconstitution of olfactory receptors that recognize overlapping sets of odorants.

The vertebrate olfactory system discriminates a wide variety of odorants by relaying coded information from olfactory sensory neurons in the olfactory epithelium to olfactory cortical areas of the brain. Recent studies have shown that the first step in odor discrimination is mediated by approximately 1000 distinct olfactory receptors, which comprise the largest family of G-protein-coupled receptors. In the present study, we used Ca(2+) imaging and single-cell reverse transcription-PCR techniques to identify mouse olfactory neurons responding to an odorant and subsequently to clone a receptor gene from the responsive cell. The functionally cloned receptors were expressed in heterologous systems, demonstrating that structurally related olfactory receptors recognized overlapping sets of odorants with distinct affinities and specificities. Our results provide direct evidence for the existence of a receptor code in which the identities of different odorants are specified by distinct combinations of odorant receptors that possess unique molecular receptive ranges. We further demonstrate that the receptor code for an odorant changes with odorant concentration. Finally, we show that odorant receptors in human embryonic kidney 293 cells couple to stimulatory G-proteins such as Galphaolf, resulting in odorant-dependent increases in cAMP. Odor discrimination is thus determined by differences in the receptive ranges of the odorant receptors that together encode specific odorant molecules.

Why are some patients with severe neglect able to copy a cube? The significance of verbal intelligence.

Cube-copying is often used to assess constructional ability of brain-damaged patients and the influence of unilateral spatial neglect is often pointed out in patients with right hemisphere lesions. However, some patients with severe neglect perform cube-copying satisfactorily. The aim of the present study is to identify the factors that affect the performance of cube-copying in patients with left unilateral spatial neglect. Constructional performance was investigated in 100 patients with unilateral spatial neglect using a task to copy the Necker cube. The relationship of the patients' cube-copying performance to the severity of their neglect, as well as other factors (verbal intelligence, age, duration after onset of the disease, educational level, lesion site, piecemeal approach, and side of starting to copy) was analyzed. Twenty-two normal subjects also participated in this study as controls. Among many factors adopted for analysis, neglect severity and verbal intelligence were found to be primary factors affecting the cube-copying performance of the patients with unilateral spatial neglect. The effect of neglect severity on cube-copying performance was apparent in the patients whose verbal intelligence was deteriorated, but was not observed in the patients with preserved verbal intelligence. Similarly, the effect of verbal intelligence on cube-copying performance was apparent in the patients with severe neglect, but not in the patients with mild neglect. We conclude that constructional ability in the copying of a cube is determined by verbal intelligence, as well as by the severity of unilateral spatial neglect.

Dynamin and rab5 regulate GRK2-dependent internalization of dopamine D2 receptors.

Dopamine D2 receptors (D2Rs; short form, which is one of the alternative splicing variants) expressed in COS-7 cells are internalized in an agonist-dependent manner only when G protein-coupled receptor kinase 2 (GRK2) is coexpressed [Ito, K., Haga, T., Lameh, J. & Sadée, W., (1999) Eur. J. Biochem. 260, 112-119]. We have examined the effects of coexpression of dynamin, a small molecular mass GTP-binding protein, rab5A, and their mutants on the internalization of D2Rs in the presence of both dopamine (10 or 100 microM) and GRK2. The rate and extent of D2R internalization was increased or decreased by coexpression of dynamin I or a dominant-negative form of dynamin I (dynamin I K44E), respectively. The effects of coexpressing these two dynamins were more prominent at 10 microM dopamine than at 100 microM. In the presence of 10 microM dopamine, internalization of D2R was completely suppressed when dynamin I K44E was coexpressed, and the half-life (t 1/2) of D2R internalization decreased relative to cells not expressing dynamin from 82 to 29 min when dynamin I was coexpressed. Internalization of D2Rs was facilitated or suppressed by coexpression of a constitutively active form of rab5A (rab5A Q79L) or a dominant-negative form of rab5A (rab5A S34N), respectively. The t 1/2 of D2R internalization at 10 microM dopamine decreased from 82 to 16 min in cells coexpressing rab5A Q79L. The effect of coexpression of rab5A S34N was more apparent at 100 microM dopamine than at 10 microM; the t 1/2 of D2R internalization at 100 microM dopamine increased from 20 to 56 min and the proportion of internalized D2Rs after 120 min decreased from 53 to 28%. These results indicate that the internalization of D2Rs is dependent on the action of dynamin as well as GRK2, and is regulated by the action of rab5A.

Functional identification and reconstitution of an odorant receptor in single olfactory neurons.

The olfactory system is remarkable in its capacity to discriminate a wide range of odorants through a series of transduction events initiated in olfactory receptor neurons. Each olfactory neuron is expected to express only a single odorant receptor gene that belongs to the G protein coupled receptor family. The ligand-receptor interaction, however, has not been clearly characterized. This study demonstrates the functional identification of olfactory receptor(s) for specific odorant(s) from single olfactory neurons by a combination of Ca2+-imaging and reverse transcription-coupled PCR analysis. First, a candidate odorant receptor was cloned from a single tissue-printed olfactory neuron that displayed odorant-induced Ca2+ increase. Next, recombinant adenovirus-mediated expression of the isolated receptor gene was established in the olfactory epithelium by using green fluorescent protein as a marker. The infected neurons elicited external Ca2+ entry when exposed to the odorant that originally was used to identify the receptor gene. Experiments performed to determine ligand specificity revealed that the odorant receptor recognized specific structural motifs within odorant molecules. The odorant receptor-mediated signal transduction appears to be reconstituted by this two-step approach: the receptor screening for given odorant(s) from single neurons and the functional expression of the receptor via recombinant adenovirus. The present approach should enable us to examine not only ligand specificity of an odorant receptor but also receptor specificity and diversity for a particular odorant of interest.

Effects of unilateral spatial neglect on spatial agraphia of kana and kanji letters.

Left unilateral spatial neglect (USN) and spatial agraphia are both caused by right hemisphere damage. However, effects of USN on spatial agraphia have not been fully investigated. This study examined performances of patients with or without neglect and normals on single letter writing, using Japanese unique writing systems, kana (phonogram) and kanji (ideogram). Our patients exhibited agraphia of kanji, but not of kana. However, accuracy of writing was not associated to neglect severity, and errors for writing to dictation occurred either on the left or right side of kanji. Right frontal damage seemed to be related to the genesis of duplication or omission errors of repeated elements. Effect of USN was limited to spatial arrangement of letters and copying performance.

[A comparative study between the clinical and autoptic findings in cases with malignant tumor in the head and neck].

In 106 autopsies of patients in Kitasato University Hospital from 1974 to 1989, who had died of malignant tumor in the head and neck, the relationship between the clinical diagnosis and its autoptic finding was retrospectively investigated. The result was summarized as follows. 1. Characteristics in 98 cases other than malignant lymphoma: 1) Local recurrence was detected by autopsy in 60 cases (61%) among 98. Discrepancy between clinical diagnosis and autoptic findings was recognized in 10 cases (10%), in whom 4 false positive cases by clinical diagnosis were included and it was considered that the rate was relatively high. 2) The metastatic lymph nodes were demonstrated by autopsy in 50 cases (51%). Discrepancy between clinical diagnosis and autoptic findings was found in 37 (38%), 35 of them were clinically diagnosed to be negative. However, in 35 of them it was considered that the discrepancy had not influenced their fatal courses. 3) Metastases to distant organs were demonstrated by autopsy in 49 (50%). Discrepancy between clinical diagnosis and autoptic findings was found in 39 (40%). 33 of them were clinically diagnosed to be negative. The discrepancy concerned with the causes of death was shown in 5 cases. 2. Characteristics in 8 cases of malignant lymphoma: Discrepancy between clinical diagnosis and autoptic findings of distant metastases was frequently observed but the discrepancy concerned with the causes of death was not shown. There were 2 cases in which side effect of chemotherapy may cause poor prognosis.

[Histological study on gland tissue in the median region of the lingual root--morphology of the foramen cecum and posterior lingual gland].

The lingual root is an area where the thyroid gland develops and small salivary glands composed of both serous and mucous glands are distributed in the submucosa. In this paper the foramen cecum and mucous glands at the root of the tongue were histologically investigated in serial sagittal sections of the medial portion of the lingual root. The materials were obtained from 59 autopsied cases, excluding the cases with metabolic disease, brain tumor and neck tumor in whom the original disease might exert direct or indirect effects to the gland tissue. The frontal end was determined to be the line which connects the right and left palatoglossal arch junctions with the tongue, and the rear end the line of transition of the vallecula to the epiglottis. The specimen was cut in half along the median lingual sulcus and median glossoepiglottic fold. Each serial section of 4 microns in thickness was cut from the median plain to the lateral and stained with Hematoxylin-Eosin or Pas-Alcian blue at pH 2.5. The results of the study were summarized as follows. 1) The foramen cecum was histopathologically confirmed in 12 cases out of 59 (20%). The ratio of the detection did not show any difference between both sexes and in the groups of different age. 2) The mucous gland opening into the mucosal epithelium, which were located at the side of the lingual apex from the Ebner's gland or serous gland, were detected in 45 out of 59 (76%).(ABSTRACT TRUNCATED AT 250 WORDS)

[Autoshaping of a button-push response and eye movement in human subjects].

Two experiments were conducted with human subjects to investigate the similarities and differences between animal and human behaviors under autoshaping procedures. In these experiments, light served as CS, and display on TV served as US. Whether the pushing button response or gazing response to CS could be obtained in human subjects under Pavlovian conditioning procedure was examined. In Experiment 1, uninstructed naive subjects were placed in a room containing a push-button and a TV display. Within the experimental sessions, the push-button was lit for 8 s as CS, and then paired with the display of a soft pornographic program on TV for 10 s. The result indicated that the modeling of pushing button promoted the increase of response probability among the subjects. The trials conducted after the rest period indicated an increase of response probability. In Experiment 2, a 4 cm square translucent panel was lit for 20 s as CS, and then paired with the display of a computer graphic picture on TV for 8 s as US. Some subjects started gazing at the CS for several seconds. These results indicated that some subjects could acquire the gazing response under the autoshaping procedure.

Recovery from drug-induced alpha coma.

A 41-year-old man in deep coma after drug overdosage showed electroencephalographic activity of alpha frequency. He remained comatose for 20 hours, and assisted ventilation was required for 28 hours, but he eventually recovered. Alpha coma has generally been considered to have grave prognostic implications, but acute drug intoxication is an exception to this rule. Intensive supportive therapy is important.