Contractions of specific abdominal muscles in postural tasks are affected by respiratory maneuvers

Hodges, Paul W., Simon C. Gandevia, and Carolyn A. Richardson. Contractions of specific abdominalmuscles in postural tasks are affected by respiratory maneuvers.J. Appl. Physiol. 83(3): 753-760, 1997.The influence of respiratory activity of the abdominal muscleson their reaction time in a postural task was evaluated. Theelectromyographic (EMG) onsets of the abdominal muscles and deltoidwere evaluated in response to shoulder flexion initiated by a visualstimulus occurring at random throughout the respiratory cycle.Increased activity of the abdominal muscles was produced by inspiratoryloading, forced expiration below functional residual capacity, and astatic glottis-closed expulsive maneuver. During quiet breathing, thelatency between activation of the abdominal muscles and deltoid was notinfluenced by the respiratory cycle. When respiratory activity of theabdominal muscles increased, the EMG onset of transversus abdominis andinternal oblique, relative to deltoid, was significantly earlier formovements beginning in expiration, compared with inspiration [by97-107 ms (P < 0.01) and64-90 ms (P < 0.01),respectively]. However, the onset of transversus abdominis EMGwas delayed by 31-54 ms (P < 0.01) when movement was performed during a static expulsive effort,compared with quiet respiration. Thus changes occur in earlyanticipatory contraction of transversus abdominis during respiratorytasks but they cannot be explained simply by existing activation of themotoneuron pool.

     

Respiration and Mitochondrial Activity in Zea mays Roots As Affected by Osmotic Stress

Studies were made of metabolism in highly vacuolated and slightlyvacuolated Zea mays root tissue both during and after plasmolysis. Plasmolysis resulted in decreased respiration and carbon dioxideevolution from glucose and an increased sucrose synthesis. Inhibitionof respiration during plasmolysis in both the highly vacuolatedand slightly vacuolated tissue was not relieved by supply ofglucose, organic acids, or uncouplers of oxidative phosphorylation.Mitochondria isolated from plasmolysed tissue were tightly coupled,but activity in vitro was inhibited by exposure to a high negativeosmotic potential. It is suggested that low TCA cycle activityin vivo must be due either to inhibition of mitochondrial activityor to reduced flow of carbon through the glycolytic pathway. A low potential for TCA cycle activity after deplasmolysis issuggested, as addition of pyruvate stimulated carbon dioxideevolution but not oxygen uptake, which was severely decreased.This was presumably due to severe mitochondrial damage as shownby their activity in vitro. However, it is not clear whetherrespiration in vivo is rate limited by rapid leakage of metabolicintermediate (reported earlier) or by lysis of mitochondria. Deplasmolysis did not damage mitochondria from slightly vacuolatedtissue, a result which was consistent with respiratory measurementsmade in vivo. The data show that mitochondria in vacuolated tissue are damagedduring and after deplasmolysis and not before. It is suggestedthat lysis of mitochondria occurs in vivo as a result of a sharpincrease in the osmotic potential of the cell fluids.     

Activity of medullary respiratory neurons during ventilator-induced apnea in sleep and wakefulness

Mechanical ventilation of cats in sleep andwakefulness causes apnea, often within two to three cycles of theventilator. We recorded 137 medullary respiratory neurons in four adultcats during eupnea and during apnea caused by mechanical ventilation. We hypothesized that the residual activity of respiratory neurons during apnea might reveal its cause(s). The results showed that residual activity depended on 1) theamount of nonrespiratory inputs to the cell (cells with morenonrespiratory inputs had greater amounts of residual activity);2) the cell type (expiratory cellshad more residual activity than inspiratory cells); and 3) the state of consciousness (moreresidual activity in wakefulness and rapid-eye-movement sleep than innon-rapid-eye-movement sleep). None of the cells showed an activationduring ventilation that could explain the apnea. Residual activity ofapproximately one-half of the cells was modulated in phase with theventilator. The strength of this modulation was quantified by using aneffect-size statistic and was found to be weak. The patterns ofmodulation did not support the idea that mechanoreceptors excite somerespiratory cells that, in turn, inhibit others. Indeed, most cells,inspiratory and expiratory, discharged during the deflation-inflationtransition of ventilation. Residual activity failed to reveal the causeof apnea but showed that during apnea respiratory neurons act as ifthey were disinhibited and disfacilitated.

     

Acclimation of Respiratory Properties of Leaves of Spinacia oleracea L., a Sun Species, and of Alocasia macrorrhiza (L.) G. Don., a Shade Species, to Changes in Growth Irradiance

To clarify the way in which the light available for growth affectsrespiration in leaves of sun and shade plants, we examined therespiratory properties of mature leaves of Spinacia oleraceaL., a sun species, and of Alocasia macrorrhiza (L.) G. Don.,a shade species, that had been grown at various irradiances.In leaves of S. oleracea, the respiratory rates, on a dry massbasis, decreased with time during the night, and the higherwas the growth irradiance during the day, the higher was therespiratory rate. The marked decreases in the respiratory rateduring the night were accompanied by decreases in the concentrationof carbohydrates in the leaves. By contrast, the respiratoryrates of leaves of A. macrorrhiza were virtually constant throughoutthe night and the absolute rates were lower than those of S.oleracea even though the absolute value of the concentrationof carbohydrates and its decrease at night resembled to thosein S. oleracea. The maximum activities of respiratory enzymeswere also similar to those in S. oleracea. However, the leavesof A. macrorrhiza contained less soluble protein than thoseof S. oleracea. These results suggest that, in S. oleracea,the concentration of carbohydrates might determine the respiratoryrate while such is not the case in A. macrorrhiza. The lowerrespiratory rates in A. macrorrhiza might be due to a lowerdemand for ATP. (Received November 29, 1995; Accepted February 15, 1996)     

PHYSIOLOGICAL.STUDIES OF A CHIMAERA FORMED BY GRAFTING NIGHTSHADE UPON TOMATO I. RESPIRATORY ACTIVITY OF LEAVES

1. Leaves of a chimaera which are composed of the tissues fromLycopersicum esculentum var. kinnari (L. e.) covered by twolayers of cells from Solatium villosum (S. v.) were comparedwith those of control plants, L. e. and S. v. with respect tothe respiratory activity. 2. As for the respiratory activity of leaf strips, the chimaerawas more active than either L. e.. or S. v. 3. To respiratory inhibitors examined leaf strips of the chimaeraresponded rather similarly to those of S. v. but differentlyfrom those of L. e. 4. Of various substrates tested, glycolic, ascorbic and chlorogenicacids were actively oxidized by cell-free extracts of leavesfrom every plant. The first two were especially good substratesfor S. v.: L. e. attacked most the third, and the chimaera oxidizedjust as well the first two as S. v. did and the third as L.e. did. 5. Ascorbic acid and chlorogenic acid contents of the leaf extractswere not correlated with the activities of the extracts of oxidizingthese substances. 6. Physiological interactions were suggested between the twotissues of different origins in the chimaera. (Received December 2, 1963; )     

Physiological Diversity in Metabolism in Marine and Terrestrial Crustacea

Two aspects of metabolic adaptation to increased terrestrialismare considered: (1) respiratory adaptations as reflected bycomparative cytochrome c oxidase activity in tissues of crabsfrom aquatic and terrestrial habitats, and (2) thermal acclimationpatterns in cytochrome c oxidase activity in tissues from thesecrabs. Enzymatic assays were done spectrophotometrically ongill, muscle, and mid-gut gland tissues from two aquatic species,Libinia emarginata and Callinectes sapidus, and the terrestrialOcypode quadrata. Cytochrome c oxidase was chosen for this studysince it is generally believed that the more aerobic the cellsor tissues become, the more fully developed the cytochrome systemwill be. This enzyme is also thought to have a role in thermalacclimation. In gill tissue the activity of cytochrome c oxidase is enhancedwith the advent of aerial respiration. Enzymatic activity ofgill tissue from Ocypode quadrata was significantly greaterthan it was in tissue from the aquatic species. No correlationwas observed with increased terrestrialism and enzymatic activityof muscle or mid-gut gland tissue. The thermal acclimation patternsof tissues of these three species of crabs indicate a clear-cuttendency for less enzymatic adaptation to temperature at thetissue level as these crabs evolve toward a land habitat.     

Changes in photosynthetic activity of the diatom Phaeodactylum tricornutum in a culture of limited volume

Changes in photosynthetic activity of a marine diatom duringalgal growth were studied with a typical culture medium formarine algae, ASP-2 (l). As the algal specimen, Phaeodactylumtricornutum was used for the experiments. Nitzschia closteriumand Chaetoceros sp. were also supplementarily used. Photosynthetic and p-benzoquinone Hill activities remarkablychanged with time during algalgrowth; with maximum activityfound in cells at log phase. A rapid decrease occurred in theinterphase from the log to stationary phase. The activity changewas not accompanied by variation in photosynthetic pigment content. The low concentration of phosphorus source was suggested asthe main cause for the change. On supply of extra inorganicphosphate, the time length for holding high photosynthetic activitybecame longer; or, the activity of the cells at stationary phaserecovered at least partly even in the dark. Dark recovery wasnot accompanied by either algal growth or an increase in thecontent of photosynthetic pigments. Inactivation of photosynthesis in the stationary growth phaseand activation by added phosphate in the dark were inferredto be due to changes in activities of both the CO₂-fixing andelectron transfer systems. The observed activity change maynot be attributable to a deficiency in inorganic phosphate asthe substrate for photophosphorylation. Similar changes in photosynthetic activity were also observedwith Nitzschia closterium and Chaetoceros sp. (Received January 30, 1971; )     

Effects of Deltamethrin on Field Grown Potato Plants:Biochemical and Ultrastructural Aspects

The effects of repeated spraying of field grown plants of Solanumtuberosum L. with deltamethrin 24, 41 and 55 d after emergence,on chlorophyll level, oxygen uptake, RuBisCo activity and leafultrastructure, were studied. Chlorophyll amount increased in treated plants after the secondand third spraying, whereas RuBisCo activity showed a highervalue than in the controls only after the last treatment. Incontrast, the respiratory activity was stimulated by the firstspraying and no significant differences were found with theensuing treatments. The general ultrastructure of mesophyllcells was not altered by deltamethrin, but morphometric analysisof electron micrographs showed that the fractional volume ofchloroplasts and starch in the treated leaves were consistentlylower than in controls. The biochemical and ultrastructural data suggested that thevegetative cycle of the sprayed plants had been extended.Copyright1993, 1999 Academic Press Solanum tuberosum L., potato plant, deltamethrin     

Development of the Recalcitrant (Homoiohydrous) Seeds of Avicennia marina: Anatomical, Ultrastructural and Biochemical Events Associated with Development from Histodifferentiation to Maturation

Early histodifferentiation of the embryo of Avicennia marina(Forssk) Vierh was characterized by the formation of endospermhaustoria Once the growth phase was initiated, subsequent embryodevelopment was extra-ovular The mature seed, therefore, wasenclosed by a pericarp originating entirely from the ovary wallGrowth and reserve deposition was not initiated until 45–50d after fruit set (DAFS), when respiratory activity had peakedWater content remained constant from the earliest stages ofembryogenesis to seed abscission and respiratory activity, althoughdeclining somewhat after the completion of histodifferentiation,remained relatively high throughout seed development The ultrastructureof the meristematic root primordia was indicative of metabolicactivity, remaining essentially similar in all respects fromthe end of histodifferentiation until the mature seeds wereabscised During this phase cotyledon cells became highly vacuolatedand the soluble sugars, which constituted the major nutrientreserves of mature seeds, increased considerably Seeds of A,marina initiate germination, without the requirement for additionalwater, as soon as they are shed It is proposed that the accumulationof soluble sugars, rather than insoluble complex reserves, isa major factor in the developmental strategy of these highlyrecalcitrant seeds Anatomy, Avicennia marina, biochemistry, homoiohydrous, recalcitrant, seed development/maturation, ultrastructure     

Catalase formation in Rhodotorula mucilaginosa in relation to the effect of glucose and antibiotics

The developmental pattern of the catalase activity in Rhodotorulamucilaginosa, an obligate aerobe, was investigated in relationto its growth. The pattern of catalase activity does not runin a manner comparable with that of a respiratory capacity,because catalase activity takes a continual rise after the middleof the logarithmic growth, while a respiratory pattern runsa constant level during the corresponding growth phase. Additionof antimycin A to cells with a minimum catalase activity doesnot block the increase in the catalase activity. Chloramphenicoldoes not exert any recognizable effect on the catalase formationwhereas cycloheximide does create an intense inhibitory effect,regardless of addition times on the course of growth. Theseresults show that the synthesizing sites of yeast catalase aredifferent from mitochondria. ¹Present address: Department of Biology, Japan Women's University,Tokyo, Japan (Received February 18, 1970; )     

The Viability and Biosynthetic Activity of Cells of Capsicum frutescens Mill. cv. annuum Immobilized in Reticulate Polyurethane

Cells of C. frutescens Mill. cv. annuum, immobilized in reticulatepolyurethane foam, retained a high level of viability aftera 23 d period of entrapment, as determined by respiratory activity,esterase activity (staining with fluorescein diacetate), nutrientuptake and secondary metabolic activity (capsaicin production).Esterase activity and the ability to accumulate capsaicin wereretained during prolonged culture periods (of up to 12 weeks'duration), and although certain precursors to capsaicin (phenylalanine,ferulic acid and isocapric acid), when supplied to immobilizedcells at a concentration of 1.0 mol m ^–3 were found toreduce respiratory activity, this could be restored by the sequentialchemical treatment of the cells, using precursor-free medium.The ability of immobilized pepper cells to produce capsaicinunder such conditions was examined. Evidence is presented whichsuggests that the immobilization in polyurethane acts to stabilizecells (in terms of the retention of esterase, respiratory andsecondary metabolic activity) when subjected to storage at 3°C. Key words: Immobilization, Viability, Secondary metabolism     

Leptin attenuates respiratory complications associated with the obese phenotype

A profile of respiratory complications has beenassociated with the onset and development of obesity in humans. Similarphenotypes have been routinely demonstrated in genetic animal models ofobesity such as the ob mouse(C57BL/6J-Lep^ob).The objective of the present study was to test the hypothesis that aconstellation of respiratory complications are attenuated with leptin(i.e., protein product of the ob gene)replacement. Daily leptin administration during a 6-wk period wasconducted to control body weight of mutantob mice similar to genotypic controlgroups. During the treatment period, repeated baseline ventilatorymeasurements were assessed by using whole body plethysmography whilequasistatic pressure-volume curves were performed to further explorethe role of leptin in improving lung mechanics. Diaphragmatic myosinheavy chain (MHC) isoform phenotype was examined to determine proportional changes in MHC composition. In room air, breathing frequency and minute ventilation were significantly(P < 0.01) different amongob treatment groups, suggesting thatleptin opposed the development of a rapid breathing pattern observed invehicle-treated ob mice. Quasistaticdeflation curves indicated that the lung volume of leptin-treatedob mice was significantly(P < 0.05) greater relative tovehicle-treated ob mice at airwaypressures between 0 and 30 cmH₂O.Diaphragm MHC composition of leptin-treated ob mice was restored significantly(P < 0.05) to resemble the controlphenotype. In this genetic mouse model of obesity, the resultssuggested that respiratory complications associated with the obesephenotype, including rapid breathing pattern at baseline, diminishedlung compliance, and abnormal respiratory muscle adaptations, areattenuated with prolonged leptin treatment.

     

Cell membrane permeability and respiratory activity in chilling stressed callus

To improve our understanding of the mechanism of chilling injuryin chill-sensitive callus (Cornus stolontfera), early changesin cell permeability and respiratory activity were studied.Partial leakage of amino acids and an abrupt increase in permeationand oxidation of added dopamine were characteristic of chilledcallus in the late stage of chilling at 0?C (48 hr), when mostof the callus sustained severe injury. However, little or nochange in cell permeability was observed in the early stageof chilling (within 24 hr), when calli retained their viabilityfor growth after transfer to a warm temperature. These resultssuggest that changes in the cell membranes per se are by nomeans the primary step in cell injury. Temporary depressionof respiratory activity was detected soon after chilling for12 hr, but activity appeared to return to the original levelon further chilling up to 24 hr. An irreversible dysfunction,however, occurred in the respiratory system on prolonged chillingup to 48 hr. This implies that irreversible impairment of mitochondrialfunctions may not be involved in the early stage of the cellinjury. A possible relationship between these observed changesand ultrastructural changes in chilled cells is discussed. ¹Contribution No. 2153 from the Institute of Low TemperatureScience. (Received June 6, 1979; )     

Ventrolateral medullary respiratory network and a model of cough motor pattern generation

The primaryhypothesis of this study was that the cough motor pattern is produced,at least in part, by the medullary respiratory neuronal network inresponse to inputs from "cough" and pulmonary stretch receptorrelay neurons in the nucleus tractus solitarii. Computer simulations ofa distributed network model with proposed connections from the nucleustractus solitarii to ventrolateral medullary respiratory neuronsproduced coughlike inspiratory and expiratory motor patterns. Predictedresponses of various "types" of neurons (I-DRIVER, I-AUG, I-DEC,E-AUG, and E-DEC) derived from the simulations were tested in vivo.Parallel and sequential responses of functionally characterizedrespiratory-modulated neurons were monitored during fictive cough indecerebrate, paralyzed, ventilated cats. Coughlike patterns in phrenicand lumbar nerves were elicited by mechanical stimulation of theintrathoracic trachea. Altered discharge patterns were measured in mosttypes of respiratory neurons during fictive cough. The resultssupported many of the specific predictions of our cough generationmodel and suggested several revisions. The two main conclusions were asfollows: 1) TheBötzinger/rostral ventral respiratory group neurons implicated inthe generation of the eupneic pattern of breathing also participate inthe configuration of the cough motor pattern.2) This altered activity ofBötzinger/rostral ventral respiratory group neurons istransmitted to phrenic, intercostal, and abdominal motoneurons via thesame bulbospinal neurons that provide descending drive during eupnea.

     

Comparative Observations on the Polarity of Sprouting of Bulbils of Dioscorea bulbifera L. and Dioscorea alata L.

The sprouting process of aerial bulbils of D. bulbifera andD. alata was compared. In D. alata, sprouts originated normallyfrom the proximal pole, at or near to the point of abscision,and irrespective of moisture, light or orientation of the bulbil.In D. bulbifera, sprouts originated primarily in response tomoisture and were less influenced by polarity. Post-harvesttreatment with gibberellic acid delayed the sprouting of bulbilsof D. alata but not of D. bulbifera; treatment with 2-chloroethanolor ethanol promoted sprouting in both species. Physiologically,sprouting was characterized by a transient increase in respiratoryrate at the onset of, or just prior to, sprout emergence. Thepromotion of sprouting by chloroethanol was accompanied by asurge in respiratory activity. Dioscorea spp., yams, bulbil germination, polarity     

"GIGANTISM" OF CHLORELLA VULGARIS I. RELATION OF GIGANTISM TO CELL GROWTH AND DIVISION

1. The sugars which induced gigantism of Chlorella cells wereglucose,fructose, galactose, mannose, xylose and arabinose.These sugarswere utilized as respiratory substrates by thealgal cells.
2. The cellular division of Chlorella was stimulatedby glucoseand galactose, but suppressed by fructose, mannose,xylose andarabinose, while all these sugars evoked gigantism.No correlationwas found between cellular division and gigantism,
3. The photosynthetic activity of giant Chlorella varied withthesorts of sugars added. It was decreased by glucose, fructoseand mannose, but was unaffected by other sugars such as galactose,xylose and arabinose.
4. The respiratory activity of giant Chlorellacells as much higherthan that of control cells.
5. The amountsof protein-N and dry weight per unit volume of giantChlorellawere much less than those of control cells.

¹ Present address: Department of Chemistry, College of GeneralEducation, Osaka University, Toyonaka, Osaka.     

Comparison of the Respiratory Metabolism of Plantago lanceolata L. and Plantago major L.

Bryce, J. H. and ap Rees, T. 1985. Comparison of the respiratorymetabolism of Plantago lanceolata L. and Plantago major L.—J.exp. Bot. 36 1559–1565. The aim of this work was to discover if the respiratory metabolismof the roots of Plantago lanceolata L. differed from that ofthe roots of Plantago major L. Measurements of oxygen uptakeand dry weight of excised root systems during growth of seedlingsprovided evidence that the two species differed in the amountof respiration needed to support a given increase in dry weight.Excised root systems were given a 6-h pulse in [U-¹⁴C]sucrosefollowed by a 16.5-h chase in sucrose. The detailed distributionof ¹⁴C amongst the major components of the roots at the endof the pulse and the chase revealed no significant differencebetween the two species. Patterns of ¹⁴CO₂ production from [1-¹⁴C],[2-¹⁴C], [3,4-¹⁴C], and [6-¹⁴C]glucose of excised root systemsfrom plants of three ages were similar for the two species.It is suggested that there is no conclusive evidence for anysignificant inherent difference in the respiratory metabolismof the roots of the two species. Key words: ¹⁴C sugar metabolism, respiration, roots, Plantago     

Respiratory Activities, Cytochrome Composition and Cytochrome c Oxidase Subunit II Levels of Mitochondria from Alloplasmic Wheats Having Aegilops Cytoplasms

Respiratory activities, cytochrome composition and cytochromec oxidase (EC 1.9.3.1 [EC] ) subunit II (COXII) levels of isolatedmitochondria in one euplasmic and four alloplasmic lines ofwheat (Triticum aestivum), having cytoplasms of Aegilops columnaris,Ae.crassa (4x),Ae. mutica and Ae. triuncialis, which show variousgrowth abnormalities, were studied. Cytoplasm-specific variationon the respiratory activities has been revealed among the cytoplasmsof five Triticum and Aegilops species. The cyanide-resistantrespiratory pathway does not seem to contribute to the variationin activity. The low level of state 3 and cytochrome oxidaseactivities were evident in lines having Ae. columnaris and Ae.triuncialis cytoplasms, which also show growth inhibition. Thelow cytochrome oxidase activity in both cytoplasms is consistentwith the low amount of cytochrome aa₃ and the severe reductionin COXII levels. The relatively low amount of cytochrome aa₃in Ae. mutica is also consistent with the reduced level of theCOXII. On the other hand, relatively low amounts of cytochromeb (b-557) are found in the Ae. crassa cytoplasmic line. Theseresults suggest that the growth abnormalities found in the alloplasmicwheat lines are related to the variation in respiratory activitiesand cytochrome composition. In particular, growth inhibitionfound in the alloplasmic wheats having Ae. columnaris and Ae.triuncialis may be caused by the severe depression of respiratoryactivity due to the impaired cytochrome oxidase activity oftheir mitochondria. ³Present address: Department of Biochemistry, Dalhousie University,Halifax, NS, Canada B3H 4H7     

Respiratory aspects of Rhodotorula mucilaginosa

The absorption spectrum of intact cels of Rhodotorula mucilaginosaexhibits three absorption peaks for carotenoids and a Soretband for cytochrome(s). The difference spectrum between thereduced and oxidized states indicates there are components ofcytochrome b, c(c₁) and a+a₃. However, the respiratory aspectof this strain differs significantly from that of baker's yeast,Saccharomyces cerevisiae. The respiratory capacity is strikinglyenhanced in the transition from the lag to the logarithmic phaseof growth, and is accompanied with a special increase in cytochromeoxidase. Addition of TMPD (plus ascorbate) in the presence ofglucose barely affects oxygen consumption of the cells duringthis period, whereas it markedly affects consumption in oldercells. Sodium azide slightly inhibits respiration in the courseof growth, except during this limited transitional period. Azide-insensitiverespiration is also seen in the mitochondrial fraction preparedfrom cells grown to the stationary state. A similar situationis found with antimycin A. Morphological investigations were carried out electron-microscopicallyon the cells of different growth stages. (Received September 16, 1969; )     

Growth, respiration and some enzymes of three strains of Ceratocystis fimbriata

Growth, respiratory activities and electrophoretic characteristicsof phosphatase and catalase in three strains of Ceratocystisfimbriata (sweet potato strain, coffee strain and prune strain)differing in pathogenicity on sweet potato roots were investigated.There were no significant differences in either growth kineticsor respiratory activity among the strains. Potassium cyanideand antimycin A inhibited oxygen uptake in sweet potato andprune strains. The oxygen uptake of endoconidia of coffee strainwas stimulated by these inhibitors. Mitochondria were preparedfrom endoconidia and mycelia of each strain, and enzyme activitiesof the electron transport system were measured. NADH₂: cytochromec oxidoreductase activity of coffee strain was higher than thatof the other strains. The electrophoretic phosphatase patternof coffee strain was identical with that of sweet potato strain,but differed from that of prune strain. On the other hand, thecatalase zymogram from prune strain was closely related to thatof sweet potato strain, but not to that of coffee strain. ¹This paper constitutes part 79 of the phytopathological chemistryof sweet potato with black rot and injury. (Received May 22, 1969; )