This paper reviews work on olfactory function performed in the author's laboratory over the last 10 years. The following aspects of this work are covered.
Neocortical olfactory areas were studied in old world monkeys. Olfactory responses were found in the lateroposterior and centroposterior portions of the orbitofrontal cortex (LPOF and CPOF). The routes of the olfactory nerve pathways to the LPOF and the CPOF were examined. An olfactory pathway to the lateral hypothalamic area (LHA) was also studied. Using unanesthetized monkeys, information processing of odors was studied in the OB, PPF-MA, LPOF, MDmc, CPOF, and LHA. In the LPOF and LHA, half or more of the cells responded differentially to one odor. We have thus been able to clearly demonstrate discrimination of odors at the cell level in these areas.

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The responses of the vascular-interstitial-lymph system to saline loading were examined in rats by continuous monitoring of circulating blood volume and simultaneous collection of thoracic duct lymph. About 90% of the infused volume shifted out of the intravascular space, of which 72% was imbibed in the interstitial space, 21% was drained through the thoracic duct, and 7% was excreted in the urine by the 50th min after the infusion amounting to 4% of body weight. The drainage of lymph protein following the infusion was about 50% of that expected from the interstitial protein concentration and even distribution of the infused saline. This result can be explained partly by the increase of lymph from skin and muscle after the infusion.

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The dependency of fluid secretion on extracellular Na+ and the levels of phosphorus compounds were studied in the perfused canine mandibular gland (using ³¹P-NMR). During control perfusion, the resting levels of creatine phosphate (CP) and ATP were 0.62±0.05 mmol•kg^-1 gland and 0.42±0.04mmol•kg^-1 (mean±S.E., n=9), respectively. Acetylcholine (Ach; 1μmol•l^-1 for 3min) induced a salivary secretion and decreased the CP level. When Na+ in the perfusate was completely replaced with Li+, Ach induced only a minimal salivary secretion and no change in the ATP and CP levels. Restitution of Na+ to the perfusion, even without added Ach, caused a decrease in ATP and CP, and a small increase in salivary secretion. These results suggest that the activity of Na+/K+ ATPase is increased inversely via a rise of the intracellular Na+ concentration and that the salivary secretion is induced not only by added secretagogues but by an increase in the Na+ entry without added secretagogues.

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The effects of changing the intracellular Ca concentration ([Ca]_i) on the calcium current (i_Ca) were studied in isolated single ventricular cells of the guinea-pig. [Ca]_i was varied by an intracellular perfusion technique using a suction pipette. i_Ca measured from internally perfused cells at a pCa lower than 9.0 was dependent on the extracellular Ca concentration ([Ca]_o). Increasing [Ca]_o from 1.8 to 5.4mM increased the amplitude of i_Ca, and reduction of [Ca]_o from 1.8 to 0.01mM decreased the amplitude. The inactivation time course of i_Ca became faster as [Ca]_o was increased and slower as [Ca]_o was reduced. By decreasing the pCa of the internal perfusate from 9.0 to 6.8, the amplitude of i_Ca was decreased markedly, but no significant change was observed in its time course. Analysis of the I-V curve led to the conclusion that a change in the driving force was not a major factor in the reduction of i_Ca. The "steady state inactivation" of i_Ca was measured by a double-pulse method. The amplitude of i_Ca elicited by the test pulse was smaller at pCa 7.4 than at pCa 9.0 at potentials of between -27 and +33mV. By normalizing the i_Ca amplitude, however, the "steady state inactivation" curve in the control and at high [Ca]_i overlapped. Similar results were obtained in Sr-Tyrode solution. The degree of "steady state inactivation" of i_Ca at the potentials positive to +10mV was larger in Ca-Tyrode than in Sr-Tyrode solution. It is proposed that the reduction in amplitude of i_Ca at higher [Ca]_i is caused by a reduction of the maximum conductance of i_Ca (g_Ca) and that Ca ions passing through i_Ca channels have a remarkable effect on its inactivation.

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The function of the pressure-overload left ventricle was studied in relation to the chamber geometry of the isolated canine heart. The occurrence of concentric hypertrophy was confirmed in dogs with aortic constriction 40 weeks after operation. The effect of the concentric hypertrophy on left ventricular function was then studied in five pressure-overload dogs. The end-diastolic pressure of the preparation was preset at 20mmHg. In the pressure-overload ventricle, the end-diastolic volume was smaller by 47% (p<0.01) and the peak systolic pressure was higher by 24% (p<0.01) than the control at isovolumic beats. Ejection pressure was controlled by our afterload controlling system at various levels and kept constant during the ejection phase. When ejection pressure was the same, the difference in stroke volume between the experimental and control groups was not significant. In the pressure-overload ventricle, the slope (the reciprocal is E_max) of the regression line of the end-systolic volume on ejection pressure decreased (p<0.01) to 0.095ml/mmHg by 47% from 0.186ml/mmHg in the control. The volume-axis intercept (V_d) of the line was reduced to 55% of the control. A decrease in the regression line slope was in linear proportion to the degree of concentric hypertrophy, namely, to the change in chamber geometry. These results indicated that: (1) Despite reduction of the end-diastolic volume, the concentric hypertrophied ventricle showed no reduction in stroke volume at the same level of ejection pressure, because of its improved capability to generate pressure. (2) E_max was dependent on ventricular geometry.

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The effects of low Na on the tetanic contractility after trains of twitch were investigated in frog skeletal muscle. Tetanic tension was induced by a stimulation of 100Hz for 400 msec every 5min and a train of twitches (150 or 250) with 1Hz was interposed between two successive tetani. When the Na concentration was reduced to 50% by replacement with choline, each twitch constituting the positive staircase during stimulus train was augmented as compared with those in normal Ringer. The amplitude of action potential during the staircase was smaller than that in normal Ringer and was gradually decreased, suggesting that low Na solution made the release of twitch-Ca from the sarcoplasmic reticulum in twitch more effective. Although the tetanic contractility in low Na was almost the same as that in normal Ringer before applying twitch train, it was more markedly inhibited after the train. The relative size of the inhibited tetanus had a correlation with the relative size of the potentiated twitch in both normal Ringer and low Na. This tetanus inhibition after the twitch train might be due to the suppression of action potential and/or due to some inhibitory factor in the excitation-contraction (E-C) coupling process.

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The distribution kinetics of an extracellular volume tracer ⁵¹Cr-EDTA was analysed from the results of continuous monitoring of plasma radioactivity. ⁵¹Cr-EDTA was injected intravenously into spleno-nephrectomized dogs by a single injection, and the radioactivity of circulating blood and physiological parameters were continuously monitored for 150min. The dilution curve of radioactivity was resolved into 4 components, and expressed as c (t)= Ae^-αt+Be^-βt+Ce^-γt+D. The half times of each term were 0.51, 2.41, and 19.26min, respectively. Based on this result, ⁵¹Cr-EDTA space was divided into 3 compartments -plasma volume (V₅₁₀), rapidly diffusing interstitial fluid (V¹), and slowly diffusing interstitial fluid (V₂). To explain the distribution of the tracer among these fluid spaces, two possible models were proposed. In the "tandem model" the volume of each compartment was calculated as 59.2, 121.7, and 101.7ml/kg of body weight, respectively, and the mobility index of the tracer from V₀ to V₁ was 5.1 times as great as that from V₁ to V₂. In the "parallel model" the volumes were 59.2, 72.3, and 151.1ml/kg of body weight and the mobility index of the tracer from V₀ to V₁ was 3.3 times as great as that from V₀ to V₂. The characteristics of each model were discussed in relation to the properties of the interstitial space.

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The effect of forskolin and isobutyl-methylxanthine on amylase release and cyclic AMP accumulation by α- and β-adrenergic agonists was studied in rat parotid slices. A small increase in cyclic AMP by β-adrenergic agonists was sufficient for maximum stimulation of amylase release (YOSHIMURA et al., 1982a), but a similar increase in cyclic AMP by forskolin did not stimulate the maximum amount of amylase release. The amount of amylase release and cyclic AMP changed in parallel when lower doses of isobutyl-methylxanthine were used, but higher doses of isobutyl-methylxanthine further increased cyclic AMP without causing a significant increase in amylase release. Amylase release stimulated to its maximum by isobutyl-methylxanthine was much lower than that by isoproterenol. These results suggest that cyclic AMP is not the only chemical correlate for the activation of amylase release by β-adrenergic agonists. The effect of phenylephrine and methoxamine on amylase release was augmented by either forskolin or isobutyl-methylxanthine, but the effect of methacholine was not. Phenylephrine increased the cyclic AMP concentration under the same conditions, but methoxamine did not. The inhibition of the effect of phenylephrine plus forskolin on cyclic AMP accumulation by propranolol was almost complete and stereospecific, but the inhibition of their effects on amylase release was incomplete and not stereospecific. Synergism of amylase release was observed in the effect between methox-amine and dibutyryl-cyclic AMP. These results suggest that the augmentation of the effect of α-adrenergic agonists on amylase release by forskolin or isobutyl-methylxanthine cannot be explained only on changes in cyclic AMP and that some other factor in collaboration with cyclic AMP may participate in the regulation of amylase release by α-adrenergic agonists.

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Comparisons were made for the ability of D-xylose to evoke changes in the transepithelial potential difference upon its addition to the mucosal solution between guinea pig and rat small intestines. Also, Na⁺-dependence of D-xylose influx from the mucosal solution into epithelial cells, and the phlorizin-sensitivity of the influx, were compared between these animals. In guinea pig intestine, the sugar caused an immediate and sustained increase in the transmural potential difference, and this potential increment was completely abolished by phlorizin. In contrast, the sugar did not cause any electrical changes in the rat small intestine. The influx of D-xylose across the mucosal border was Na⁺-dependent and phlorizin-sensitive in guinea pigs, while it was Na⁺-independent and phlorizin-insensitive in rats. The results indicate that there is a qualitative difference in the mode of transport of D-xylase between guinea pigs and rats.

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We have recently proposed that the total mechanical energy generated in each cardiac contraction can be quantified by the systolic pressure-volume area (PVA). PVA is the area in the pressure-volume (P-V) diagram that is circumscribed by the end-systolic and end-diastolic P-V relation curves and the systolic segment of the P-V trajectory. This area has dimensions of energy and comprises the external mechanical work and the elastic potential energy. In the left ventricle of cross-circulated canine hearts, we studied the relation between PVA and oxygen consumption per beat (V_o2) above V_o2for mechanically unloaded contraction. We assumed that this excess V_o2is utilized for mechanical contraction by the contractile machinery. The percentage of PVA in the excess V_o2, both in the same unit of energy, J, would then represent the efficiency of energy conversion from the excess V_o2to the total mechanical energyin the contractile machinery. We obtained this efficiency in variously loaded contractions in both control and enhanced contractile states with epinephrine and calcium. We found that the efficiency was constant at 30-50 (mean 40) % regardless of the changes in both loading conditions and contractile states. By this constant efficiency and a variable fraction of external work in PVA, we accounted for the load- and contractility-dependent variability of the conventional mechanical efficiency (0-30%) of the heart.

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The isolated longitudinal or circular muscle strip of rat uterus on day 20 of pregnancy becomes quiescent when incubated with Krebs solution for about 2hr. Electric stimulation gave rise to a twitch-like contraction with a small amplitude instead of the spontaneous phasic contraction observed during the early period of incubation. When saponin (30-500μg/ml) was applied, the basal tension was elevated and spontaneous contractions having large amplitude were elicited. The amplitude was larger than the phasic contraction of the K-contracture observed prior to the saponin treatment. The membrane was depolarized by about 25mV, and periodic burst discharge was generated at the onset of saponin treatment. Raising the external Mg concentration from 0 to 2.4mM in steps caused a depression of contractions in a dose-de-pendent manner before treatment with Saponin, whereas the depression by Mg was much reduced after saponin treatment. The saponin-treated muscle recovered to exhibit control activity when tissues were superfused with Krebs solution for hours, i.e. the membrane was repolarized, the duration of action potential was detracted, and the depressant effect of Mg was again potentiated. For comparison, the effects of saponin and Mg on the contractions of ileal longitudinal muscle of pregnant rat were studied. In view of the above observations, the following were discussed. 1. Saponin impaired the depressant effect of Mg on the generation of contractions. 2. The membrane once impaired by saponin recovered in vitro probably under the influence of the genomic effect of ovarian hormones, a phenomenon which appeared characteristic for pregnant uterus.

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To analyze the effect of changes in interstitial fluid on lymph production, intravascular infusions of saline were given to splenectomized rats under pentobarbital anesthesia at 3 different rates (2, 3, and 4ml/100g of body weight) over 10min. Change of blood volume was continuously monitored and, simultaneously, thoracic duct lymph was collected during and after the infusion. Equilibrium was attained approximately 40min after the infusion; regardless of the infusion rate, 10% of the infused volume was incorporated into the vascular space and 90% was filtered into the extravascular space. Thus, the amount of transvascular fluid shift showed a linear relationship with the infused volume. However, the drainage from thoracic duct lymph amounted to 5.9%, 11.4%, and 17.8% of the infused saline volume when given at the rate of 2, 3, and 4ml/100g, respectively. The relation of lymph flow and infused volume could be regarded as a nonlinear system. By means of a simulation study, this relation was found to be attributed to the nonlinearity of conductance for fluid movement from tissue to lymph duct, which was only one sixth of that determined for the capillary membrane. The drainage of lymph protein following the infusion was only about 50% of that expected from the interstitial protein concentration in even distribution of the infused saline. These characteristics of interstitial fluid space play an important role in absorbing water and, hence, buffering changes in circulating blood volume after volume loading.

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Chromaffin cells isolated from bovine adrenals and maintained in culture were loaded with ⁴⁵Ca, and the mechanism of ⁴⁵Ca efflux from these cells was investigated. During exposure to a Ca-deficient, Mg (1mM)-containing medium, replacement of NaCl with sucrose, choline Cl, or Tris Cl reduced the rate of ⁴⁵Ca efflux by approx. 30-40%, showing the presence of a Na-dependent ⁴⁵Ca efflux mechanism. In contrast to the previous findings, however, Li can partially replace Na in maintaining ⁴⁵Ca efflux. Reintroduction of Ca to a Ca-deficient medium increased the rate of ⁴⁵Ca efflux depending on the concentrations of Ca, showing the presence of a Ca-dependent Ca efflux mechanism. The marked but transient rise in the rate of ⁴⁵Ca efflux in response to Ca reintroduction was observed under conditions in which the concentration of internal Na was raised either by inhibiting Na pumping activity (ouabain or K removal) or by removal of Mg from the medium. The increased ⁴⁵Ca efflux was further potentiated by simultaneous Na removal and was markedly inhibited by Ca channel blockers, suggesting that the transiently increased Ca influx in exchange for internal Na facilitated the rate of ⁴⁵Ca efflux. However, other conditions which evoke catecholamine secretion by stimulating Ca influx also increased the rate of ⁴⁵Ca efflux, suggesting that the increased Ca influx via any route increases 45Ca efflux. These results thus confirmed the presence of both Na-dependent and Ca-dependent Ca efflux mechanisms in chromaffin cells.

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Firing neuron R14 in the Aplysia parietovisceral ganglion enhances the force of stimulated contractions and causes rhythmic contractions rather than individual contractions in response to a single stimulation of the anterior aorta of Aplysia. Bath application of 1.0mM glycine to the artery causes a small depolarization, rhythmic contractions and enhances the force of individual, neurally induced contractions of the anterior aorta. These observations suggest that the physiological role of the innervation of the anterior aorta by R14 is to convert the mode of contraction of the anterior aorta from direct neural control to a myogenic mode. R14 activity does not produce classical junction potentials and inotophoretically applied glycine does not produce detectable potential changes in the anterior aorta muscle fibers. Glycine increases the force and frequency of Aplysia heart beat, and blood pressure. Glycine causes release of preloaded Ca²⁺ from the ventricle of the Aplysiaheart, implying that glycine causes intracellular release of Ca²⁺. The similarity of the actions of glycine and electrical activity in R14 extend previous evidence that R14 may utilize glycine to modulate the contractility of Aplysia circulatory muscle.

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Reactive hyperemia in the calf after arresting femoral blood flow for 3min was measured by means of strain gauge plethysmography in different postures, i.e. supine, sitting with legs resting horizontally (sitting I), and sitting with the feet on the ground (sitting II). Blood flow in the calf at rest and hyperemia decreased more in the sitting I and II positions than in the supine position. However, the peak flow of the reactive hyperemia was higher in the sitting I and II positions than in the supine position.

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In voltage-clamped guinea-pig Purkinje fibers, removal of acetylcholine (ACh) in the presence of isoproterenol produced a rebound increase in the slow inward current. This effect of ACh was abolished by atropine. These results are consistent with the hypothesis that cessation of the stimulation of myocardial muscarinic receptors transiently leads to enhanced activity of the β-receptor/slow channel system in this tissue.

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