Comparative responses of tracheal spirals and parenchymal strips to histamine and carbachol in vitro. The responses of isolated guinea pig tracheal spirals and parenchymal strips to histamine and carbachol were compared. The parenchymal strip, a 1.5 x 1.5 x 20-mm strip cut from the periphery of the lung, constricted at a lower dose and had a larger maximal response to histamine than to carbachol. In contrast, the response of the tracheal spiral to equimolar doses of histamine or carbachol was the same. The responsiveness of both muscle strips to histamine was decreased by treatment with the H1 receptor antagonist mepyramine (0.1 micrometer), and the response to carbachol was blocked by treatment with atropine (0.1 micrometer). Indomethacin (3 micrometer), cimetidine (1 micrometer), propranolol (10 micrometer), and N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) (4 micrometer) did not alter the differential response of the two strips to histamine and carbachol. The differential response of parenchymal strips with many, few, or no conducting airways and blood vessels was identical, suggesting that the contractile element is alveolar duct smooth muscle or alveolar contractile elements. This differential pharmacologic response in vitro is consistent with the in vivo observation that histamine causes more peripheral airway constriction than does acetylcholine.


Neurodiagnostic abnormalities in patients with acute renal failure. Neurological abnormalities are a major cause of morbidity in patients with renal failure. The pathophysiology of these neurological changes is unclear, and the effects on them of dialysis and return of renal function have not been well studied. Studies were done in 31 patients who had acute renal failure (ARF), all of whom were either treated with dialysis within 5 days or did not survive. Studies on these patients included the electroencephalogram (EEG), motor nerve conduction velocity, and plasma Ca(++) and parathyroid hormone (PTH) levels. Studies were done at the time ARF was diagnosed, after stabilization on dialysis, during the diuretic phase of ARF, and 3 mo after recovery from ARF. In 16 patients with acute or chronic renal failure who did not survive and in nine patients without renal disease who died, measurements were made in brain of content of Na(+), K(+), Cl(-), Ca(++), Mg(++), and water. In patients with ARF for less than 48 h, despite the fact that there were only modest increases in plasma urea and creatinine, there were striking abnormalities in the EEG. The percent EEG power < 5 Hz+/-SE was 41+/-8% (normal = 2+/-1%), whereas the percent of frequencies > 9 Hz was only 22+/-6% (normal = 62+/-3%). These changes were unaffected by dialysis, but became normal with return of renal function and remained normal at 3 mo follow-up. The motor nerve conduction velocity was unaffected by either ARF or dialysis. In patients with ARF, the brain Ca(++) was 46.5+/-3.2 meq/kg dry wt, almost twice the normal value of 26.9+/-1.0 meq/kg dry wt (P < 0.001). The plasma PTH level was 3.2+/-0.6 ng/ml (normal < 1.5 ng/ml, P < 0.01). The increased brain Ca(++) was not related to an increased plasma (Ca(++)) (PO(4) (---)) product (r(2) = 0.14, P > 0.05). There was a small but significant decrement in brain Na(+) (P < 0.05), but brain water, K(+), and Mg(++) were unaffected by ARF.Thus, in patients with ARF for less than 48 h, the EEG is grossly abnormal and there are elevated levels of PTH in plasma. The PTH appears to have a direct effect on the brain, resulting in an increased brain Ca(++) content. The EEG abnormalities are unaffected by dialysis, but they become normal with return of renal function and remain normal after 3 mo follow-up. Thus, PTH may be a major uremic toxin, demonstrating evidence for central nervous system toxicity when there are only minimal abnormalities of other biochemical markers of ARF.


Increased adrenal sensitivity to angiotensin II in low-renin essential hypertension. Studies were undertaken to determine if the dissociation of aldosterone and plasma renin activity in low-renin essential hypertension is due to altered adrenal responsiveness to angiotensin II. The responsiveness of the adrenal glands to angiotensin II was determined by infusing graded doses of angiotensin II into normal subjects and into patients with essential hypertension and measuring changes in levels of plasma aldosterone in response to the infusion. To minimize the influence of endogenous angiotensin II and ACTH, supplemental sodium and dexamethasone were given before the infusions. Levels of plasma aldosterone and plasma renin activity were determined in normal subjects and in the same patients after the combined stimuli of furosemide and upright posture, a maneuver used to increase the level of endogenous angiotensin II. To determine if the changes in levels of plasma aldosterone during infusion of angiotensin II were due to alteration of the metabolic clearance of aldosterone, the metabolic clearance of aldosterone was measured before and during the infusion of angiotensin II. After sodium loading, dexamethasone treatment, and supine posture, levels of plasma aldosterone of normal subjects and patients with essential hypertension were suppressed equally. In response to the infusion of angiotensin II, the levels of plasma aldosterone of patients with low-renin essential hypertension were significantly higher than those of normal subjects or of patients with normal-renin essential hypertension. After furosemide and upright posture, levels of plasma aldosterone of patients with low-renin essential hypertension were significantly higher than those of patients with normal-renin essential hypertension, despite a blunted response in plasma renin activity of the patients with low-renin essential hypertension. Decreases in metabolic clearance of aldosterone during infusion of angiotensin II were similar in patients with normal-renin essential hypertension and in patients with low-renin essential hypertension and accounted for only a small fraction of the marked increase in levels of plasma aldosterone of patients with low-renin essential hypertension. It is concluded that patients with low-renin essential hypertension have increased adrenal sensitivity to angiotensin II. This increased sensitivity may explain the dissociation of aldosterone and plasma renin activity in low-renin essential hypertension.


Failure of hypoxic pulmonary vasoconstriction in the canine asthma model. Effect of prostaglandin inhibitors. Measurements of respiratory mechanics, arterial blood gases, and pulmonary vascular resistance were made before and 15 min after inhalation challenge with Ascaris suum extract in dogs with natural sensitivity to this antigen. 25 of 47 dogs were treated before inhalation challenge with a prostaglandin inhibitor (90 mg/kg of aspirin or 2 mg/kg of indomethacin by intravenous infusion). In response to the challenge, bronchospasm developed in approximately half (responders) of each group reflected by decreases in mean specific respiratory system conductance and arterial oxygen tension. While the dogs were breathing room air, pulmonary vascular resistance remained unchanged after antigen challenge in the responders not given aspirin or indomethacin, but increased significantly and was associated with a lesser degree of arterial hypoxemia in the responders pretreated with either of the prostaglandin inhibitors. Prevention of arterial hypoxemia by oxygen breathing blocked an increase in pulmonary vascular resistance in four pretreated responders. No changes in respiratory mechanics, pulmonary hemodynamics, or arterial blood gases were noted in the 21 dogs who did not develop bronchospasm regardless of whether or not they were pretreated. 12 additional dogs in whom arterial hypoxemia was produced by 10% oxygen breathing, showed an increase in pulmonary vascular resistance that was not potentiated by pretreatment with aspirin in 6. We conclude that in acute experimental canine asthma, vasodilator prostaglandins appear to blunt the hypoxic pulmonary vasoconstrictor response, thereby further compromising gas exchange but preventing the development of pulmonary hypertension.


Effect of starvation on the turnover and metabolic response to leucine. l-Leucine was administered as a primed continuous 3-4-h infusion in nonobese and obese subjects in the postabsorptive state and for 12 h in obese subjects after a 3-day and 4-wk fast. In nonobese and obese subjects studied in the post-absorptive state, the leucine infusion resulted in a 150-200% rise in plasma leucine above preinfusion levels, a small decrease in plasma glucose, and unchanged levels of plasma insulin and glucagon and blood ketones. Plasma isoleucine (60-70%) and valine (35-40%) declined to a greater extent than other amino acids (P < 0.001). After 3 days and 4 wk of fasting, equimolar infusions of leucine resulted in two- to threefold greater increments in plasma leucine as compared to post-absorptive subjects, a 30-40% decline in other plasma amino acids, and a 25-30% decrease in negative nitrogen balance. Urinary excretion of 3-methylhistidine was however, unchanged. Plasma glucose which declined in 3-day fasted subjects after leucine administration, surprisingly rose by 20 mg/100 ml after 4 wk of fasting. The rise in blood glucose occurred in the absence of changes in plasma glucagon and insulin and in the face of a 15% decline in endogenous glucose production (as measured by infusion of [3-(3)H]glucose). On the other hand, fractional glucose utilization fell by 30% (P < 0.001), thereby accounting for hyperglycemia. The estimated metabolic clearance rate of leucine fell by 48% after 3 days of fasting whereas the plasma delivery rate of leucine was unchanged, thereby accounting for a 40% rise in plasma leucine during early starvation. After a 4-wk fast, the estimated metabolic clearance rate of leucine declined further to 59% below base line. Plasma leucine nevertheless fell to postabsorptive levels as the plasma delivery rate of leucine decreased 65% below postabsorptive values. (a) Infusion of exogenous leucine in prolonged fasting results in a decline in plasma levels of other amino acids, improvement in nitrogen balance and unchanged excretion of 3-methylhistidine, thus suggesting stimulation of muscle protein synthesis, (b) leucine infusion also reduces glucose production and to an even greater extent, glucose consumption, thereby raising blood glucose concentration; and (c) the rise in plasma leucine in early starvation results primarily from a decrease in leucine clearance which drops progressively during starvation.


3-Hydroxyproline content of normal urine. Values for total 3-hydroxyproline and 4-hydroxyproline were obtained from 24-h urine specimens of 18 healthy human subjects of both sexes, whose ages ranged from the first to the sixth decade in age. Urinary 3-hydroxyproline levels, not earlier described to our knowledge, were determined by an isotope-dilution method requiring considerable purification and utilizing the amino acid analyzer for final measurement. 3-Hydroxyproline averaged 3% of the corresponding 4-hydroxyproline in individual urine samples. Like 4-hydroxyproline, 3-hydroxyproline excretion is increased in the second decade, and there is generally good correlation between the two values in individual urines. A hydroxyprolinemic subject excreting greatly elevated 4-hydroxyproline levels did not excrete excessive 3-hydroxyproline, consistent with independent catabolic pathways for the two compounds. 3-Hydroxyproline appears to be selectively excreted relative to 4-hydroxyproline when compared with the probable total body content of each amino acid. Possible explanations are: a more rapid turnover of basement membrane collagen than interstitial collagen or, alternatively, relatively greater resistance to the proteolytic cleavage of peptides containing 3-hydroxyproline.


Functional profile of the isolated uremic nephron. Role of compensatory hypertrophy in the control of fluid reabsorption by the proximal straight tubule. An in vitro approach to the study of single nephron function in uremia has been employed in evaluating the control of fluid reabsorption by the renal superficial proximal straight tubule (PST). Isolated segments of PSTs from the remnant kidneys of uremic rabbits (stage III) were perfused in vitro and their rate of fluid reabsorption compared with normal PSTs and with PSTs derived from the remnant kidneys of nonuremic rabbits (stage II). All segments were exposed to a peritubular bathing medium of both normal and uremic rabbit serum thereby permitting a differentiation to be made between adaptations in function which are intrinsic to the tubular epithelium and those which are dependent upon a uremic milieu.Compared with normal and stage II PSTs, there was significant hypertrophy of the stage III tubules as evidenced by an increase in length and internal diameter, and a twofold increase in the dry weight per unit length. Fluid reabsorption per unit length of tubule was 70% greater in stage III than in normal and stage II PSTs, and was closely correlated with the increase in dry weight. Substitutions between normal and uremic rabbit serum in the peritubular bathing medium did not affect fluid reabsorption significantly in any of the three groups of PSTs. Perfusion of the tubules with an ultrafiltrate of normal vs. uremic serum likewise failed to influence the rate of net fluid reabsorption. It has previously been observed that net fluid secretion may occur in nonperfused or stop-flow perfused normal rabbit PSTs exposed to human uremic serum. Additional studies were thus performed on normal and stage III PSTs to evaluate whether net secretion occurs in the presence of rabbit uremic serum. No evidence for net secretion was found. These studies demonstrate that fluid reabsorption is greatly increased in the superficial PST of the uremic remnant kidney and that this functional adaptation is closely correlated with compensatory hypertrophy of the segment. Humoral factors in the peritubular environment do not appear to be important mediators of the enhanced fluid reabsorption.


Activation of human factor IX (Christmas factor). Human Factor IX (Christmas factor) is a single-chain plasma glycoprotein (mol wt 57,000) that participates in the middle phase of the intrinsic pathway of blood coagulation. It is present in plasma as a zymogen and is converted to a serine protease, Factor IXabeta, by Factor XIa (activated plasma thromboplastin antecedent) in the presence of calcium ions. In the activation reaction, two internal peptide bonds are hydrolyzed in Factor IX. These cleavages occur at a specific arginyl-alanine peptide bond and a specific arginyl-valine peptide bond. This results in the release of an activation peptide (mol wt approximately equal to 11,000) from the internal region of the precursor molecule and the generation of Factor IXabeta (mol wt approximately equal to 46,000). Factor IXabeta is composed of a light chain (mol wt approximately equal to 18,000) and a heavy chain (mol wt approximately equal to 28,000), and these chains are held together by a disulfide bond(s). The light chain originates from the amino terminal portion of the precursor molecule and has an amino terminal sequence of Tyr-Asn-Ser-Gly-Lys. The heavy chain originates from the carboxyl terminal region of the precursor molecule and contains an amino terminal sequence of Val-Val-Gly-Gly-Glu. The heavy chain of Factor IXabeta also contains the active site sequence of Phe-Cys-Ala-Gly-Phe-His-Glu-Gly-Arg-Asp-Ser-Cys-Gln-Gly-Asp-SER-Gly-Gly-Pro. The active site serine residue is shown in capital letters. Factor IX is also converted to Factor IXaalpha by a protease from Russell's viper venom. This activation reaction, however, occurs in a single step and involves only the cleavage of the internal arginyl-valine peptide bond. Human Factor IXabeta was inhibited by human antithrombin III by the formation of a one-to-one complex of enzyme and inhibitor. In this reaction, the inhibitor was tightly bound to the heavy chain of the enzyme. These data indicate that the mechanism of activation of human Factor IX and its inhibition by antithrombin III is essentially identical to that previously shown for bovine Factor IX.


Choline pathways during normal and stimulated renal growth in rats. Cellular membrane synthesis occurs during normal and stimulated renal growth. Choline in the kidney is utilized as a precursor for membrane synthesis via the choline kinase reaction. We investigated choline phosphorylation during normal and stimulated renal growth. Rapidly growing neonatal rat kidneys contained relatively high levels of choline kinase activity (61 pmol phosphorylcholine/min per mg protein). Choline kinase activity and phosphorylcholine production then fell gradually over the 1st mo of life; by 1 mo phosphorylcholine production was 34 pmol phosphorylcholine/min per mg protein. Choline kinase activity increased by 27% (P less than 0.001) in 28-day-old rats when renal growth was stimulated by contralateral nephrectomy; the increase occurred within 2 h after surgery. Thus, changes in the activity of this important enzyme in the initiation of membrane synthesis is associated both with normal renal development and with adaptation to nephron loss. The findings further suggest that the cell membrane may be involved in the initiation of compensatory renal growth.


Differentiation of macrophages from normal human bone marrow in liquid culture. Electron microscopy and cytochemistry. To study the various stages of human mononuclear phagocyte maturation, we cultivated bone marrow in an in vitro diffusion chamber with the cells growing in suspension and upon a dialysis membrane. At 2, 7, and 14 days, the cultured cells were examined by electron microscopy and cytochemical techniques for peroxidase and for more limited analysis of acid phosphatase and arylsulfatase. Peroxidase was being synthesized in promonocytes of 2- and 7-day cultures, as evidenced by reaction product in the rough-surfaced endoplasmic reticulum, Golgi complex, and storage granules. Peroxidase synthesis had ceased in monocytes and the enzyme appeared only in some granules. By 7 days, large macrophages predominated, containing numerous peroxidase-positive storage granules, and heterophagy of dying cells was evident. By 14 days, the most prevalent cell type was the large peroxidase-negative macrophage. Thus, peroxidase is present in high concentrations in immature cells but absent at later stages, presumably a result of degranulation of peroxidase-positive storage granules. Clusters of peroxidase-negative macrophages with indistinct borders (epithelioid cells), as well as obvious multinucleated giant cells, were noted. Frequently, the interdigitating plasma membranes of neighboring macrophages showed a modification resembling a septate junction--to our knowledge, representing the first documentation of this specialized cell contact between normal macrophages. We suggest that such junctions may serve as zones of adhesion between epithelioid cells.


Isolation of circulating immune complexes using Raji cells. Separation of antigens from immune complexes and production of antiserum. Raji cells were used for the isolation of complement-fixing antigen-antibody complexes from serum. Immune complexes bound to these cells were radiolabeled at the cell surface with lactoperoxidase. The complexes were then eluted from the cells with isotonic citrate buffer pH 3.2 or recovered by immunoprecipitation of cell lysates. The antigen and antibody moieties of the complexes were isolated by dissociating sucrose density gradient centrifugation or by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A variety of preformed immune complexes were successfully isolated from serum with this approach. In addition, these techniques were used to isolate and identify the antigens in immune complexes in the serum of rabbits with chronic serum sickness and rats with Moloney virus-induced sarcomas. Methods were also developed for the production of antisera against the antigenic moiety of immune complexes isolated from serum. Repeated challenge of rabbits with whole Raji cells with bound complexes or eluates from such cells resulted in antibody production against the antigens of the immune complexes, although reactivity against cellular and serum components was also elicited. Monospecific antisera against the antigens in immune complexes were produced by immunizing rabbits with the alum-precipitated antigen isolated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These techniques may be useful in isolating antigens in immune complex-associated diseases of unknown etiology.


Control of the myocardial contractile state by carotid chemo- and baroreceptor and pulmonary inflation reflexes in conscious dogs. The effects of carotid chemoreceptor stimulation with intracarotid injections of either nicotine, 0.2 mug/kg, or cyanide, 2 mug/kg, were compared with the effects of bilateral carotid occlusion on left ventricular (LV) pressure, dP/dt, and diameter in conscious dogs instrumented with ultrasonic diameter gauges and miniature pressure gauges. With heart rate maintained constant, carotid chemoreceptor stimulation increased mean arterial pressure by 27+/-3%, LV and diastolic diameter by 4+/-0.9% and LV dP/dt by 21+/-2%. With ventilation controlled during succinylcholine infusion, carotid chemoreceptor stimulation increased mean arterial pressure by 43+/-2% and dP/dt by 37+/-5%, values significantly greater, P < 0.01, than were observed in dogs with spontaneous ventilation. Similarly, in dogs with spontaneous ventilation after vagotomy, carotid chemoreceptor stimulation also increased dP/dt by a greater amount, i.e., by 48+/-9%. The increases in LV end diastolic diameter were not affected significantly by either cholinergic blockade with atropine or beta adrenergic blockade with propranolol. Although cholinergic blockade did not affect the inotropic or pressor responses significantly, beta adrenergic blockade attenuated the pressor response and essentially abolished the inotropic response. Bilateral carotid occlusion increased mean arterial pressure and LV end diastolic diameter by similar amounts to those observed with chemoreceptor stimulation, but increased dP/dt significantly less, P < 0.02, i.e., by 13+/-2%. As was observed with chemoreceptor stimulation, inotropic responses were not affected significantly by cholinergic blockade, but were essentially abolished by beta adrenergic blockade. Thus, in the conscious dog with heart rate constant, carotid chemoreceptor stimulation induces a clear positive inotropic effect, which is greater in the absence of the attenuating influences of pulmonary inflation reflexes, and for an equal elevation in arterial pressure appears to exert a greater increase in myocardial contractility than does carotid baro-receptor unloading.


Correction of cobalamin malabsorption in pancreatic insufficiency with a cobalamin analogue that binds with high affinity to R protein but not to intrinsic factor. In vivo evidence that a failure to partially degrade R protein is responsible for cobalamin malabsorption in pancreatic insufficiency. In vitro studies indicate that [(57)Co]cobalamin (Cbl) is preferentially bound to salivary R protein as opposed to intrinsic factor (IF) and that [(57)Co]Cbl bound to R protein is not transferred to IF at either pH 2 or pH 8. Incubation of R protein-[(57)Co]Cbl with pancreatic proteases causes a partial degradation of the R protein moiety and a rapid transfer of [(57)Co]Cbl to IF. We have postulated that the etiology of Cbl malabsorption in pancreatic insufficiency is an inability to partially degrade R protein because of a lack of pancreatic proteases. We have tested this hypothesis by determining the ability of a nonradioactive Cbl analogue, bound with high affinity by R protein but not by IF, to correct the malabsorption of [(57)Co]Cbl in patients with pancreatic insufficiency.R protein bound the Cbl analogue known as cobinamide with affinities that were the same and only 14-fold lower than those for Cbl at pH 8 and pH 2, respectively. Cobinamide was bound by IF with affinities that were 600,000- and 10,000-fold lower than those for Cbl at pH 8 and 2, respectively. The addition of 125 pmol of nonradioactive cobinamide to 0.5 pmol of [(57)Co]Cbl before being added to 1 pmol of R protein and 1 pmol of IF, markedly inhibited the ability of R protein to compete with IF for binding the [(57)Co]Cbl. Similar results were obtained with freshly aspirated gastric juice. This change was essentially indistinguishable from that observed previously when R protein or R protein-[(57)Co]Cbl was incubated in vitro with trypsin. The oral administration of 100 nmol of nonradioactive cobinamide in Schilling tests was equivalent to trypsin in its ability to completely correct the malabsorption of 0.4 nmol of [(57)Co]Cbl in three patients with pancreatic insufficiency. The fact that both trypsin and nonradioactive cobinamide inhibit the ability of R protein to compete with IF for [(57)Co]Cbl binding in vitro, and correct the mal-absorption of [(57)Co]Cbl in patients with pancreatic insufficiency in vivo, supports our hypothesis that the primary defect in Cbl absorption in this disease is an inability to partially degrade R protein because of a lack of pancreatic proteases.


Plasmid-mediated resistance to antibiotic synergism in enterococci. Mating experiments have shown that high-level resistance (minimal inhibitory concentration greater than 2,000 microgram/ml) to streptomycin and kanamycin, and resistance to penicillin-streptomycin and penicillin-kanamycin synergism are transferable by conjugation from resistant clinical isolates of enterococci to a sensitive recipient strain. Cesium chloride-ethidium bromide ultracentrifugation revealed a satellite (plasmid) band in resistant clinical isolates and the transconjugant strains but not in the sensitive recipient. Examination of these satellite bands by agarose gel electrophoresis and electron microscopy demonstrated a common plasmid with a weight of 45 megadaltons. Novobiocin treatment of a resistant clinical isolate produced simultaneous loss of high-level resistance to streptomycin and kanamycin, and of resistance to penicillin-aminoglycoside synergism. These results suggest that (a) high-level resistance to streptomycin and kanamycin among some clinical isolates of enterococci is associated with a 45 megadalton plasmid, and (b) the same plasmid is also responsible for the resistance to penicillin-aminoglycoside synergism observed in these strains.


Spironolactone. An aldosterone agonist in the stimulation of H+ secretion by turtle urinary bladder. In the isolated turtle bladder, spironolactone inhibits sodium transport in the presence of aldosterone or endogenous mineralocorticoid hormone. In contrast to this antagonism for the stimulation of sodium transport by aldosterone, the stimulation of hydrogen ion secretion by aldosterone is not inhibited by spironolactone. In hormone-depleted bladders, spironolactone stimulates hydrogen ion secretion. The extent of stimulation is similar to that of aldosterone. Spironolactone functions as an agonist for aldosterone for the stimulation of urinary acidification.


Methoscopolamine inhibition of sleep-related growth hormone secretion. Evidence for a cholinergic secretory mechanism. We have examined the effects of cholinergic blockade with 0.5 mg methscopolamine bromide, intramuscularly, on sleep-related and insulin-induced growth hormone (GH) secretion. 17 normal young men were studied; 8 had sleep studies, and 12 (including 3 who also had sleep studies) had insulin tolerance tests (ITT) with 0.1 U/kg of regular insulin. After an adjustment night in the sleep laboratory, saline control night and methscopolamine night studies were done in random sequence; study procedures included electroencephalographic, electromyographic, and electrooculographic recordings, and blood sampling every 20 min for hormone radioimmunoassays. Prolactin levels were also measured during sleep. For methscopolamine night studies, the mean overall control GH level of 2.89+/-0.44 ng/ml and the mean peak control GH level of 11.09+/-3.11 ng/ml were dramatically reduced to 0.75+/-0.01 and 1.04+/-0.25 ng/ml, respectively (P<0.0001 and <0.001). Despite virtual absence of GH secretion during the night in every study subject, no measured sleep characteristic was affected by methscopolamine, including total slow-wave sleep (12.1+/-2.6% control vs. 10.3+/-2.5% drug, P>0.2). Sleep prolactin levels were not changed by methscopolamine. In contrast to the abolition of sleep-related GH secretion, administration of methscopolamine had only a marginal effect on the GH response to insulin hypoglycemia. None of nine time points differed significantly, as was also the case with peak levels, mean increments, and areas under the curves (P>0.2). Analysis of variance did, however, indicate that the lower GH concentrations achieved during ITT after methscopolamine (average 31.7% below control) were significantly different than control concentrations. We conclude that the burst of GH secretion which normally occurs after sleep onset is primed by a cholinergic mechanism which does not influence slow-wave sleep. Cholinergic mechanisms do not appear to play an important role in sleep-related prolactin secretion. The contrast between the complete suppression of sleep-related GH release and the relatively small inhibitory effect on ITT-induced GH secretion suggests that the neurotransmitter mechanisms, and presumably the pathways, which subserve sleep-related GH secretion in man may be different from those which mediate the GH response to pharmacologic stimuli such as insulin.


Lithium inhibition of bone mineralization and osteoid formation. Lithium chloride administration to growing rats, which resulted in circulating lithium levels of 1.4 meq/liter, was attended by significant suppression of bone mineralization and organic matrix synthesis as assessed by tetracycline labeling and histological quantitation of osteoid, respectively. These effects of lithium were not associated with changes in animal behavior, nor were there any significant differences in blood levels of calcium, phosphorus, alkaline phosphatase, creatinine, pH, or parathyroid hormone. The data suggest that lithium inhibition of bone mineralization is secondary to suppression of osteoid formation.


Synthesis of DNA and poly(adenosine diphosphate ribose) in normal and chronic lymphocytic leukemia lymphocytes. Peripheral blood lymphocytes were isolated from 9 patients with chronic lymphocytic leukemia (CLL) and 12 normal control donors. The cells were assayed for synthesis of DNA and poly-(adenosine diphosphate ribose) (poly[ADPR]) immediately after isolation and on successive days following their treatment with phytohemagglutinin (PHA). Two different techniques were used to measure DNA synthesis. In the standard technique, DNA synthesis was measured by incubating intact cells with [(3)H]deoxythymidine. In the new technique, the lymphocytes were first rendered permeable to nucleotides, then DNA synthesis was measured by incubating them with [(3)H]deoxythymidine triphosphate in the presence of deoxyATP, deoxyGTP, deoxyCTP, ATP, and Mg(++). Both assays showed the anticipated rise in DNA synthesis after PHA stimulation of normal cells. PHA-stimulated lymphocytes from patients with CLL demonstrated low levels of DNA synthesis in both assay systems. The initial levels of poly(ADPR) synthesis were greater in CLL lymphocytes than in normal cells. Studies with a T-cell-depleted population of normal cells showed the same activity for poly(ADPR) synthesis that was demonstrated by the original population of normal cells. PHA stimulation produced an increase in poly(ADPR) synthesis in both the normal and CLL cells. The increase in poly(ADPR) synthesis in normal cells was coincident with the increase in DNA synthesis. The increase in poly(ADPR) synthesis in the CLL cells was dissociated from the delayed and diminished increase in DNA synthesis. Thus, CLL cells have higher than normal initial levels of poly(ADPR) synthesis. Poly(ADPR) synthesis is dissociated from DNA synthesis in CLL cells whereas it varies directly with DNA synthesis in normal lymphocytes.


Extrapancreatic glucagon and glucagonlike immunoreactivity in depancreatized dogs. A quantitative assessment of secretion rates and anatomical delineation of sources. The anatomical sites and the rates of extrapancreatic secretion of glucagon and of glucagon-like immunoreactivity (GLI) were assessed in dogs 2 h after pancreatectomy by catheterization of the gastrosplenic and mesenteric veins. Glucagon release from the gastrosplenic area approximated one-fourth that of a normal pancreas and rose from 0.25 to 1.0 ng/kg per min during arginine stimulation. Intestinal glucagon secretion was small and did not respond to arginine, suggesting that the stomach is the only important extrapancreatic source of glucagon. Glucagon concentrations attained by gastrosplenic secretion were in close proportion to those obtained during the administration of exogenous glucagon, indicating similar clearance rates of extrapancreatic and pancreatic glucagon, approximating 10 ml/kg per min.GLI secretion (0.3 ng eq/kg per min) was limited to the intestinal area and was transiently stimulated by arginine and exogenous glucagon. Base-line GLI clearance approximated 1 ml/kg per min. No insulin secretion could be detected. Gastrointestinal glucose uptake rose from 0.56 to 2.2 mg/kg per min after glucagon administration suggesting that as much as 10% of total glucose production can be taken up by the gastrointestinal tract. In two dogs both the stomach and pancreas were removed. Intestinal glucagon release remained small and did not increase during arginine administration. By contrast, GLI release was stimulated by both arginine and exogenous glucagon.


Regulation of blood flow to the aortic media in dogs. Morphologic observations suggest that the inner layers of the thoracic aorta in man and dog are avascular and the outer layers have vasa vasorum. It appears that vasa vasorum are essential in the thoracic aorta because their interruption produces medial necrosis. These experiments provide the first measurements of blood flow through aortic vasa vasorum and examine physiologic regulation of that flow. During control conditions the outer two-thirds of the media of the thoracic aorta received 10 ml/min per 100 g blood flow through vasa vasorum. Flow to the inner third of the aorta was 1 ml/min per 100 g. Flow to both the inner and outer media of the abdominal aorta was less than 1 ml/min per 100 g. Adenosine increased blood flow to vasa vasorum in the outer media of the thoracic aorta from 7 to 18 ml/min per 100 g, but did not increase flow to the inner layers of the aorta. Hemorrhagic hypotension decreased flow in the outer media of the thoracic aorta from 14 to 2 ml/min per 100 g. Acute hypertension failed to increase blood flow through vasa vasorum, as conductance decreased significantly. These studies indicate that vasa vasorum provide a considerable amount of blood flow to the outer layers of the thoracic aorta. The vessels are responsive to physiologic stimuli because they dilate during infusion of adenosine and constrict during both hemorrhagic hypotension and acute hypertension. We speculate that the failure of blood flow to the aortic wall to increase during acute hypertension might, if it were sustained, contribute to aortic medial necrosis.


Indomethacin is a placental vasodilator in the dog. The effect of prostaglandin inhibition. The effect of 8 mg/kg of indomethacin on uterine blood flow, prostaglandin production, and intraamniotic fluid pressure was examined in late pregnant dogs. Uterine blood flow was measured with 15 mum radiolabeled microspheres. Because we found that a significant percentage of the microspheres shunted through the placental circulation into the lungs, we calculated placental blood flow by adding the shunted microspheres through the placenta to the nonshunted microspheres in the placenta. Total uterine blood flow significantly increased from 271+/-69 ml/min during control period to 371+/-72 ml/min (P < 0.01) 30 min after indomethacin. This increase was attributable to the change in blood flow to the placental circulation (222+/-58 to 325+/-63 ml/min; P < 0.01). Associated with these hemodynamic changes we found an almost complete suppression of uterine prostaglandin E(2) production (1,654+/-305 to 51+/-25 pg/ml; P < 0.01) as measured by gas chromatography-mass spectrometry. In addition, we found that indomethacin treatment resulted in uterine relaxation as measured by intraamniotic fluid pressure changes (11.2+/-1.3 mm Hg to 8.5+/-1.2 mm Hg; P < 0.001). We conclude that indomethacin causes an increase in placental blood flow without any change in flow to the rest of the uterus, and that this dose of the drug inhibits greater than 95% of uterine prostaglandin production. In addition, indomethacin is responsible for uterine relaxation. The increase in placental blood flow after indomethacin is probably a result of uterine relaxation, which is secondary to prostaglandin synthesis inhibition.


Metabolism of cholesterol-rich chylomicroms. Mechanism of binding and uptake of cholesteryl esters by the vascular bed of the perfused rat heart. The rate of uptake of cholesteryl ester from chylomicrons has been determined with the isolated perfused rat heart and both intact and functionally hepatectomized rats. Uptake was found to be proportional to the cholesteryl ester content of the particles. Transfer of cholesteryl ester to other lipoprotein classes of the plasma was negligible under these conditions, and loss of cholesteryl ester from the medium was associated with quantitative recovery in the vascular bed. The uptake mechanism was nonsaturable and independent of the lipoprotein lipase binding site. Compared with receptor-dependent uptake of low density lipoprotein cholesteryl ester by heart endothelium, the chylomicron pathway appears to provide a major proportion of cholesteryl ester cleared from the plasma. Uptake was initially heparin dependent, and cleared lipid was released by 10 microgram/ml of heparin; however, lipid taken up rapidly became heparin resistant and was then hydrolyzed slowly with production of unesterified fatty acid. These results are discussed in the context of the possible role of cholesterol-rich chylomicron remnant lipoproteins in atherogenesis.


Oral glucose augmentation of insulin secretion. Interactions of gastric inhibitory polypeptide with ambient glucose and insulin levels. Gastric inhibitory polypeptide, or GIP, has been postulated as the major enteric hormonal mediator of insulin release. The release of immuno-reactive GIP (IR-GIP) after oral glucose and its role in insulin release was studied in normal men by the glucose clamp technique. In 24 subjects studied with the hyperglycemic clamp, blood glucose was maintained at 125 mg/dl above basal for 2 h via a primed-continuous IV glucose infusion coupled to a servo-controlled negative feedback system. 40 g glucose per m(2) surface area was ingested at 60 min, and the blood glucose was maintained at the steady-state hyperglycemic level. Plasma IR-GIP and insulin (IRI) levels were measured throughout the 2-h period. IR-GIP levels changed little when IV glucose alone was given; the mean basal value was 305+/-34 (SEM) pg/ml. After oral glucose, IR-GIP levels began to rise within 10 min and reached a peak within 40 min of 752+/-105 pg/ml. Plasma IRI responded initially to the square wave of hyperglycemia in the typical biphasic pattern. After oral glucose, plasma IRI levels rose strikingly above the elevated levels produced by hyperglycemia alone, reaching a peak of 170+/-15 muU/ml within 45 min. The time course of the rise in IR-GIP and IRI was nearly identical. To assess whether the maintenance of euglycemia would affect this process, the euglycemic clamp was employed in 11 subjects to maintain basal blood glucose levels during a similar 2-h study. A primed-continuous insulin infusion, with a constant rate of 120 mU/m(2) per min was given together with a servo-controlled glucose infusion. This resulted in hyper-insulinemia of approximately 300 muU/ml. Glucose was ingested by six subjects at 60 min. Plasma IR-GIP responded to oral glucose similarly to the effect seen in the hyperglycemic studies. No increase in endogenous insulin release was seen despite the increase in IR-GIP when euglycemia was maintained. However, in five of seven subjects given insulin whose blood glucose concentration rose by 20 mg/dl or more after oral glucose, there was an increase in plasma insulin concentration associated with the elevation in IR-GIP. Thus, the effect of glucose-released IR-GIP on insulin secretion is dependent upon the presence of some degree of hyper-glycemia and is not inhibited in the presence of marked hyperinsulinemia.


Factors influencing the handling of insulin by the isolated rat kidney. The renal handling of immunoreactive insulin was studied in the isolated perfused normothermic rat kidney to determine (a) the relative contributions of glomerular clearance and peritubular clearance to the renal clearance of insulin under different conditions, (b) what metabolic factors influence the ability of tubular cells to remove insulin from the glomerular filtrate and the peritubular circulation, and (c) whether the same factors influence the luminal and contraluminal uptake of insulin.In control kidneys the organ clearance of insulin (OCi) was 974+/-63 mul/min (SEM), of which a maximum of 46% could theoretically be accounted for by filtration. OCi was not altered by fasting, lack of exogenous fuel (glucose), or the addition of cyanide. The glomerular filtration rate did not correlate with the OCi, but there was a significant (P < 0.001) negative correlation (r = -0.828) between the peritubular clearance and glomerular filtration rate. Both N-ethylmaleimide and cold (10 degrees C) reduced the rate of insulin removal. Fractional excretion of filtered insulin (9.7+/-1.7% in controls) was not significantly altered by fasting or perfusing without glucose. In contrast, KCN increased fractional excretion of insulin to 41.9+/-3.7% whereas cold increased fractional excretion to 69.0+/-3.3%. This study indicates that renal tubular cells remove insulin from the tubular lumen and the peritubular compartment. Furthermore, the data suggest that insulin removal by tubular cells is a temperature-sensitive process consisting of two different systems. The system associated with the luminal aspect of the cell appears to be dependent on oxidative metabolism, whereas the system associated with the contraluminal aspects of the cell appears to be independent thereof. Under several circumstances when the glomerular clearance of insulin falls thereby reducing the amount of insulin absorbed by the luminal aspect of the cell, contraluminal uptake increases, and a constant rate of insulin removal is maintained by the kidney.


IgE receptors on human basophils. Relationship to serum IgE concentration. As reported previously, and confirmed here in 26 donors, the serum IgE level (2.6-5,500 ng/ml) correlates well (rs = 0.95, P less than 0.001) with the in vivo number of IgE molecules/basophil (6,000-600,000). The total number of IgE receptors/basophil was monitored by incubating them with an IgE-rich serum (15 microgram/ml), quantitatively stripping IgE from the cells at pH 3.7, and measuring eluted IgE by a direct radioimmunosorbent test. Saturation of receptors for each donor was achieved with 15 nM IgE (3 microgram/ml). The proportion of receptors occupied in vivo correlated with the serum IgE (rs = 0.84, P less than 0.001) whereas the average association constant of the receptors was independent of serum IgE and ranged from 7.1 X 10(8)/M to 2.8 X 10(10)/M, averaging 7.7 X 10(9)/M. Unexpectedly, the total number of IgE receptors/basophil was closely related to the serum IgE level. (rs = 0.92, P less than 0.001). Thus, either there is genetic association between serum IgE and the number of basophil IgE receptors, or, more likely, the receptor number is modulated by the serum IgE concentration.


Neutralization of glucagon by antiserum as a tool in glucagon physiology. Lack of depression of basal blood glucose after antiserum treatment in rats. The method of producing experimental glucagon deficiency by administration of glucagon antiserum was evaluated in rats. A pool of antisera was prepared, the affinity of which exceeded that of the glucagon receptors of liver cell membranes, whereas the binding capacity of the volume used amounted to more than one-third of the total glucagon content in the rat pancreas. That rapid, extensive, and lasting neutralization of glucagon had taken place after antiserum treatment was indicated by the following findings: When examined more than 1 h after the injection and after 60 min of exercise-stimulated glucagon production, all rats had excess free antibodies in plasma. The concentration of free glucagon was lowered to one-third of the concentration in control rats; at 37 degrees C plasma samples could bind 25% of additional 300 pmol/liter of glucagon in 10 s, and 69% in 120 s; the glycemic response to exogenous glucagon was abolished. Antiserum treatment, however, had no effect on blood glucose in rats fasted for 3 and 10 h, in chemically sympathectomized and adrenomedullectomized rats, and in 48-h-fasted, acutely adrenalectomized rats. The antiserum was found to contain 460 nmol/liter of antibody-bound glucagon, originating in the rabbit in which the antiserum was raised. However, antibody preparations from which the bound glucagon had been effectively removed were equally ineffective in lowering the basal blood glucose in rats, although in three-fourths of the rats the concentration of free glucagon was lowered beyond detection limit. The data indicate that the absolute concentration of glucagon in plasma is of minor importance for the maintenance of basal blood glucose in the rat.


Kinetics of human connecting peptide in normal and diabetic subjects. The metabolic clearance rate (MCR) of synthetic human connecting peptide (C-peptide) was measured with a single-dose injection technique in six normal and seven diabetic subjects and with a constant infusion technique in one normal subject. The MCR of C-peptide did not differ in normal subjects (4.4 ml/min per kg; range, 3.7-4.9) and in diabetic subjects (4.7 ml/min per kg; range, 3.7-5.8). Employment of both techniques in one subject gave similar MCR. The average half-life of C-peptide in plasma calculated from the last 1-h period of the single-dose injection studies was longer in the insulin-dependent diabetics (42.5 min; range, 39.4-48.5) than in the normal subjects (33.5 min; range, 24.9-45.3). These results indicate that the beta-cell secretory capacity of normal and insulin-dependent diabetic subjects can be compared by measuring the C-peptide concentration in peripheral venous plasma. The difference in the half-life of C-peptide in plasma between diabetics and normals suggests an altered kinetics of the disappearance of the peptide, while the overall metabolism, as expressed by the MCR, is similar.


Abnormal carbohydrate metabolism in chronic renal failure. The potential role of accelerated glucose production, increased gluconeogenesis, and impaired glucose disposal. To delineate the potential role of disordered glucose and glucose-precursor kinetics in the abnormal carbohydrate metabolism of chronic renal failure, alanine and glucose production and utilization and gluconeogenesis from alanine were studied in patients with chronic compensated renal insufficiency and in normal volunteers. With simultaneous primed injection-continuous infusions of radiolabeled alanine and glucose, rates of metabolite turnover and precursor-product interrelationships were calculated from the plateau portion of the appropriate specific activity curves. All subjects were studied in the postabsorption state. In 13 patients with chronic renal failure (creatinine = 10.7+/-1.2 mg/100 ml; mean+/-SEM), glucose turnover was found to be 1,035+/-99.3 mumol/min. This rate was increased 56% (P = 0.003) over that observed in control subjects (664+/-33.5 mumol/min). Alanine turnover was 474+/-96.0 mumol/min in azotemic patients. This rate was 191% greater (P = 0.007) than the rate determined in control subjects (163+/-19.4 mumol/min). Gluconeogenesis from alanine and the percent of glucose production contributed by gluconeogenesis from alanine were increased in patients with chronic renal failure (192% and 169%, respectively) as compared to controls (P < 0.05 for each). Alanine utilization for gluconeogenesis was increased from 40.2+/-3.86 mumol/min in control subjects to 143+/-39.0 mumol/min in azotemic patients (P < 0.05). The percent of alanine utilization accounted for by gluconeogenesis was not altered in chronic renal insufficiency. In nondiabetic azotemic subjects, mean fasting glucose and immunoreactive insulin levels were increased 24.3% (P = 0.005) and 130% (P = 0.046), respectively.These results in patients with chronic renal failure demonstrate (a) increased glucose production and utilization, (b) increased gluconeogenesis from alanine, (c) increased alanine production and utilization, and (d) a relative impairment to glucose disposal. We conclude that chronic azotemia is characterized by increased rates of glucose and glucose precursor flux and by a relative impairment to glucose disposal. These findings may suggest an underlying hepatic and peripheral insensitivity to the metabolic action of insulin in patients with chronic renal insufficiency.


A specific inhibitor of complement (C5)-derived chemotactic activity in serum from patients with systemic lupus erythematosus. In the course of examining polymorphonuclear leukocyte (PMN) chemotaxis in patients with systemic lupus erythematosus (SLE), we have found a previously undescribed serum inhibitor of complement (C5)-derived chemotactic activity. Serum from a 25-yr-old Black female with untreated SLE, when activated with zymosan, failed completely to attract either her own or normal PMN. Incubation of normal PMN with the patient's serum did not affect their subsequent random motility or chemotactic response toward normal zymosan-treated serum (ZTS). The patient's serum, however, did inhibit the chemotactic activity of normal ZTS and of column-purified C5-derived peptide(s), but had no effect on the chemotactic activity of either the synthetic peptide, N-formylmethionyl leucyl-phenylalanine or a filtrate prepared from a culture of Escherichia coli (bacterial chemotactic factor). The inhibitory activity in the patient's serum resisted heating at 56 degrees C for 30 min and could be separated from C5-derived chemotactic activity in the patient's ZTS (or normal ZTS that had been incubated with the patient's serum) by chromatography on Sephadex G-75. Despite its effect on C5-derived chemotactic activity, the patient's serum did not influence two other C5-derived biologic activities: PMN lysosomal enzyme-releasing activity and PMN-aggregating activity. Chromatography of the patient's serum (65% ammonium sulfate pellet) on Sephadex G-200 yielded three distinct peaks of inhibitory activity. Two were heat labile and exhibited other properties of the previously described chemotactic factor inactivators of normal human serum. The third and most active peak, however, resisted heating at 56 degrees C for 30 min, eluted with an apparent mol wt of 50,000-60,000, and acted specifically on C5-derived chemotactic activity. This uniquely specific, heat-stable inhibitor of C5-derived chemotactic activity has been found thus far in serum from 4 of 11 patients with active SLE and may account, in part, for altered host defenses against infections caused by pyogenic microorganisms.


Study of chloride transport across the rabbit cortical collecting tubule. Recent micropuncture studies have suggested that the collecting tubule may be involved in the regulation of extracellular fluid volume. The present studies were designed to evaluate chloride transport across the in vitro-perfused rabbit cortical collecting tubule inasmuch as chloride ion would ultimately affect extracellular fluid volume. The tubules were perfused and bathed with artificial solutions simulating ultrafiltrate. Four groups of studies were conducted. In groups one and two, tubules from rabbits not receiving desoxycorticosterone (DOCA) were compared to tubules from rabbits which had received DOCA (5 mg/day) for 1 wk. In groups three and four, tubules were obtained only from rabbits not receiving DOCA. In group one, sequential bidirectional chloride fluxes were measured. The ratio of chloride efflux to influx was 0.99+/-0.04 in tubules obtained from rabbits not receiving DOCA whereas it was 1.28+/-0.09 in tubules obtained from rabbits receiving DOCA, suggesting stimulation of net chloride flux under these conditions. In group 2, chemical chloride concentration and osmolality of the collected fluid were measured. Neither the chemical chloride concentration nor the osmolality of the collected fluid decreased significantly below their respective perfusion fluid values in tubules from non-DOCA-treated rabbits but there was a significant decrease in the chemical chloride concentration (10-42 meq/liter) and osmolality (10-42 mosmol/kg H(2)O of the collected fluid in tubules from DOCA-treated rabbits. In group three, unidirectional chloride permeabilities from lumen-to-bath were determined during the passage of current down the perfusion pipette. The alterations of the average lumen potential, -35+/-4 and +28+/-2 mV, did not influence unidirectional chloride movement suggesting that the cortical collecting tubule is quite impermeable to chloride. In group four, unidirectional chloride permeability from lumen-to-bath was measured before and after substitution of NaCH(3)SO(4) for sodium chloride in the bath. Replacement of chloride by CH(3)SO(4) reversibly decreased the apparent chloride permeability from 2.41+/-0.50 to 0.69+/-0.08 (x10(-5) cm/s) demonstrating that (36)Cl permeability is dependent on the chemical concentration of chloride.THE CURRENT STUDIES DEMONSTRATE THAT: (a) the cortical collecting tubule is able to reabsorb salt under the modulation of circulating mineralocorticoids and, thus, may participate in overall volume homeostasis; (b) the chloride permeability and the major portion of isotopic chloride flux across the cortical collecting tubule is via exchange diffusion; and (c) under certain circumstances the cortical collecting tubule may act as a diluting segment.


The activation of plasminogen by Hageman factor (Factor XII) and Hageman factor fragments. Activation of plasminogen through surface-mediated reactions is well recognized. In the presence of kaolin, purified Hageman factor (Factor XII) changed plasminogen to plasmin, as assayed upon a synthetic amide substrate and by fibrinolysis. Kinetic studies suggested an enzymatic action of Hageman factor upon its substrate, plasminogen. Hageman factor fragments, at a protein concentration equivalent to whole Hageman factor, activated plasminogen to a lesser extent. These protein preparations were not contaminated with other agents implicated in surface-mediated fibrinolysis. Diisopropyl fluorophosphate treatment of plasminogen did not inhibit its activation by Hageman factor. These studies indicate that Hageman factor has a hitherto unsuspected function, the direct activation of plasminogen.


Glucagon metabolism in the rat.

Glucagon metabolism in the rat. The renal handling of the biologically active glucagon component (the 3,500-mol wt fraction of immunoreactive glucagon [IRG]) and the contribution of the kidney to its overall peripheral metabolism were studied in normal and uremic rats. The metabolic clearance rate of glucagon was 31.8 +/- 1.2 ml/min per kg in normal animals and was diminished by approximately one-third in each of three groups of rats with compromized renal function: 22.3+/-1.6 ml/min per kg in partially (70%) nephrectomized; 22.9+/-3.3 ml/min per kg in bilaterally ureteral ligated; and 23.2+/-1.2 ml/min per kg in bilaterally nephrectomized animals. In normal rats the kidney contributed 30% to the overall metabolic clearance of the hormone and the renal extraction of endogenous and exogenous glucagon was similar, averaging 22.9+/-1.6% and was independent of plasma IRG levels over a wide range of arterial concentrations. The remnant kidney of partially (70%) nephrectomized animals continued to extract substantial amounts (16.6+/-4.2%) of the hormone, but accounted for only 8% of the total peripheral catabolism of IRG. In the two groups of animals with filtering kidneys, renal glucagon uptake was linearly related to its filtered load and could be accounted for by glomerular filtration and tubular reabsorption. However, the kidneys of animals with both ureters ligated (renal extraction of inulin = 3.2+/-1.8%) and hence virtual absence of glomerular filtration, continued to extract 11.5+/-1.9% of the renal arterial glucagon, contributing by 9% to its overall metabolic clearance, indicating that IRG uptake occurs also from the post glomerular capillaries.


Application of the solid phase C1q and Raji cell radioimmune assays for the detection of circulating immune complexes in glomerulonephritis. The C1q solid phase and Raji cell radioimmune assays were used to determine the frequency of detectable circulating immune complexes in patients with glomerulonephritis. In this study, 46% of 56 patients with glomerulonephritis had evidence of circulating immune complexes. More important, circulating immune complexes were associated with some, but not other, types of glomerulonephritis. Thus, immune complexes were detected in lupus glomerulonephritis (9/9 patients), rapidly progressive glomerulonephritis (5/6 patients), and acute nephritis (5/6 patients), but not in IgA-IgG glomerulonephritis (0/7 patients), or membranous glomerulonephritis (0/8 patients). The Raji cell radioimmune assay and the C1q solid phase radioimmune assay showed concordance of 79% in the detection of circulating immune complexes. Serial determinations, in general, showed either persistence of a negative or positive result of conversion of positive to negative.


Long-term regulation of adipocyte glucose transport capacity by circulating insulin in rats. We have tested the idea that the circulating plasma insulin level plays an important role in the long-term regulation, or maintenance, of the cellular glucose transport system, distinct from insulin's ability to acutely accelerate glucose transport. To study this hypothesis, groups of rats were made either hyperinsulinemic or hypoinsulinemic by daily insulin injections, or streptozotocin treatment, respectively. Different levels of hypoinsulinemia were produced by using different doses of streptozotocin (40 and 55 mg/kg). The mean (+/-SE) 9 a.m. plasma insulin level for each experimental group was: hyperinsulinemic animals, 65+/-5 muU/ml; controls, 32+/-3 muU/ml; low dose streptozotocin group, 18+/-3 muU/ml; and high dose streptozotocin group 5+/-2 muU/ml. Isolated adipocytes were prepared from each animal and glucose transport was assessed by measuring the initial rates of uptake of the nonmetabolyzable hexose 2-deoxy glucose. The V(max) and K(m) values for adipocyte glucose transport were calculated from the 2-deoxy glucose uptake data. The results demonstrated that in cells from control animals the V(max) of in vitro adipocyte glucose transport was 7.1+/-0.7 nmol/min per 10(6) cells in the basal state and 22.9+/-0.9 nmol/min per 10(6) cells in the presence of a maximally effective insulin concentration (25 ng/ml) in the buffer. In cells from the experimentally hyperinsulinemic animals these V(max) values were increased to 11.7+/-0.8 and 44.2+/-1.1 nmol/min per 10(6) cells. Using adipocytes from both groups of streptozotocin-treated (high dose, 55 mg/kg; low dose, 40 mg/kg) insulin-deficient diabetic animals, V(max) values were found to be progressively decreased. Thus, in the low dose group, basal-and insulin-stimulated V(max) values were 1.6+/-0.5 and 5.7+/-0.7 nmol/min per 10(6) cells, as compared to values of 0.9+/-0.2 and 1.7+/-0.6 in the high dose group. Thus, when considered as group data a positive relationship was found between circulating plasma insulin levels and adipocyte glucose transport V(max), with increased V(max) values in hyperinsulinemic rats and decreased V(max) values in hypoinsulinemic rats. Furthermore, when the individual data were analyzed, highly significant correlation coefficients were found between the height of the plasma insulin level and both the basal (r = 0.82, P < 0.001) and insulin-stimulated (r = 0.93, P < 0.001) V(max) values. The apparent K(m) for 2-deoxy glucose uptake was the same under all conditions. In conclusion, assuming that the V(max) of transport is some function of the number of glucose transport carriers per cell, then these results support the hypothesis that in addition to acute acceleration of glucose transport, insulin is also an important long-term regulator of the number of available adipocyte glucose transport carriers.


Viscoelastic properties of the rectal wall in Hirschsprung's disease. Viscoelastic properties of the rectal wall were compared with Hirschsprung's disease. The elasticity of the rectal wall after accomodation to distension was found to be significantly greater (P less than 0.001) in patients, and the time taken by the rectum to accomodate was also found to be longer (P less than 0.001). The increased elasticity correlated well with severity of the illness, but none of the parameters correlated with length of aganglionic segment. Measuring elastic properties of the rectal wall may help to assess the severity of illness in patients with Hirschsprung's disease.


Effect of parathyroid hormone and uremia on peripheral nerve calcium and motor nerve conduction velocity. Peripheral neuropathy is not an uncommon complication of chronic uremia. Because parathyroid hormone, by raising brain calcium, is partly responsible for central nervous system aberrations in uremia, we studied the relative role of uremia, per se, and(or) parathyroid hormone on peripheral nerve calcium and motor nerve conduction velocity (MNCV). Studies were made in six groups of six dogs each, as follows: (a) normal dogs, (b) thyroparathyroidectomized (T-PTX) animals, (c) dogs with 3 days of uremia produced by bilateral nephrectomy, (d) T-PTX before the induction of acute renal failure, (e) normal dogs receiving 100 U/day of parathyroid extract (PTE) for 3 days, and (f) normal animals receiving 3 days of PTE followed by 5 days without PTE. Calcium content in peripheral nerve (expressed as milligram per kilogram of dry weight) was 252+/-5 (SE) in normal animals and 262+/-4 in T-PTX dogs. It was significantly (P < 0.01) higher in dogs with acute renal failure and intact parathyroid glands (410+/-12) and in normal animals receiving PTE (362+/-7). T-PTX, before acute renal failure, prevented the rise in peripheral nerve calcium (262+/-4) and PTE withdrawal was followed by the return of peripheral nerve calcium to normal (261+/-3). The increments in peripheral nerve calcium were associated with slowing of MNCV. It decreased significantly from 70+/-4 to 43+/-1 m/s after 3 days of acute uremia in dogs with intact parathyroid glands and T-PTX before acute renal failure prevented the fall in MNCV. Administration of PTE to normal animals reduced MNCV from 63+/-3 to 35+/-3 m/s and the withdrawal of PTE restored MNCV to normal (73+/-2 m/s). The results show that (a) excess parathyroid hormone increases peripheral nerve calcium and slows MNCV, (b) T-PTX, previously performed, prevents these changes in acute uremia, and (c) the withdrawal of PTE administration is followed by a reversal of the abnormalities.


The nature of the immunoreactive lipotropins in human plasma and tissue extracts. This study was designed to establish definitively the nature of immunoreactive lipotropin (IR-LPH) in human plasma and tissue extracts. Using gel filtration, gel filtration under denaturing conditions, cationic exchange chromatography, immunoprecipitation, and radioimmunoassay, we have studied normal and tumorous human pituitaries, ectopic ACTH- and LPH-secreting tumors, plasma from normal subjects before and after dexamethasone administration, and plasma from patients with primary adrenal insufficiency and pituitary and nonpituitary ACTH- and LPH-secreting tumors. Except in the plasma and tumors of occasional patients with ectopic ACTH syndrome, the smallest IR-LPH appears to be lambda-lipotropin (lambdaLPH), which is often the predominant and occasionally the only IR-LPH present. The other major peptide appears to be betaLPH, a 91-amino acid molecule that contains lambdaLPH as its 1-58 sequence. Larger immunoreactive materials were observed in some specimens, but the "big" LPH in one plasma was shown to be lambdaLPH bound to IgG.The weak melanocyte-stimulating activity of LPH suggests that ACTH may be the principal pigmentary hormone in man. The fact that lambdaLPH, rather than betaLPH, is the predominant form in plasma suggests that the enkephalin-endorphin opiate peptides, which are contained in the "missing" 59-91 sequence from the betaLPH precursor of lambdaLPH, may be secreted in parallel with ACTH under both physiological and pathological conditions in man.


The perceptual use of semantic rules by normal-hearing and hard-of-hearing children. This study investigated the effects of varying degrees of linguistic constraint upon the performance of hard-of-hearing subjects in a repetition task. Experimental stimuli consisted of three sets of sentence strings: grammatical, semantically anomalous, and ungrammatical, respectively. While the control group performed essentially equally across all three conditions, the hard-of-hearing group performed best to the grammatical stimuli and worst to the ungrammatical stimuli. It was hypothesized that the hard-of-hearing subjects employed a primitive sentence dictionary, while the normal-hearing subjects had developed a more sophisticated word dictionary.


Speech development in hearing-impaired children. Many hearing-impaired children are unable to speak intelligibly. Research indicates, however, that patterns of errors typically found in the speech of such children could be largely avoided by systematic teaching. The purpose of this paper is to present and discuss a model for the progressive development of speech skills to the levels of automaticity required for fluent spoken language.


Amplification for hearing-impaired children: basic considerations. The importance of the selection of proper amplification for the hearing impaired child is discussed including goals of amplification, specific methods of choosing amplification, and test procedures including speech audiometry. Evaluation controversies discussed include: monaural versus binaural amplification; body worn versus post auricular hearing aids; maximum output; and special hearing aid modifications.


Relationship of word knowledge to word frequency in young adult deaf students. This report describes research that has been undertaken to determine the relationship between word knowledge and word frequency (occurrence in print) in a deaf population. A matching test was designed and administered to 162 deaf students entering the National Technical Institute for the Deaf in the summer of 1976. The results of the study indicate that there exists a relationship between word frequency and word knowledge in a deaf population. The reliability and validity of the technique was tested and seems to provide adequate support for this approach to assessing the vocabulary level of deaf students.


That deaf child and you: a forensic approach to the problems of hearing and speech. Gesture, particularly manual gesture, is an important element in early childhood language formation. Its attempted elimination from the early communicative experience of hearing impaired children raises serious ethical, and possibly legal, questions. Though deaf children are deprived of information by the suppression of Sign Language so that their pretended education becomes a nullity, their cognitive power remains intact and undiminished. Official and academic refusals to recognize gesture and abhorrence, explicit or implied, of non-vocal language systems are culture-specific in their operation. They have no effect on normal children but are highly destructive of hearing-impaired children in direct proportion to the degree of impairment. Thus, sign-deprivation in profound hearing loss is completely destructive. The responsibility, including legal liability, of professionals stands in the same proportion. The most important developmental period for Sign Language is infancy. It is doubly important therefore that, in addition to educators and indeed prior to them, all health professionals who work with hearing impairment of any type be proficient in the principles and practice of nonvocal communication. Sign Language is the hearing aid of the deaf.


Toward a psychology of deafness?

Toward a psychology of deafness? A large scale study comparing Yugoslavian and American adolescents with normal hearing and with early profound deafness suggests that a relative tendency to impulsivity is a notable characteristic of the deaf group. Despite the conclusiveness of the study overall, there was little intercorrelation among individual subjects on the several test measures employed. A stereotype should be avoided therefore, and the tests viewed as perhaps tapping different personality aspects or avenues of development for what clinically becomes a final common pathway. Comparable and comparative studies of deaf children of deaf parents and of children whose hearing parents have learned manual language early may define how much the tendency is based on qualities of parental discomfort with a deaf child, on the unavailability of language, or on the absence of sound alone.


Sign codes and sign language: two orders of communication. Deaf children, like all children, are born with a capacity for language; i.e. the symbolization and expression of cognitive functioning. Until the age at which hearing children are talking, deaf children communicate well with others, using their sight, touch, and other actions--the common gestures in the culture of their caretakers. The chief problem in communication arises when hearers reject the deaf child's gestural symbolization of his developing language knowledge. The problem is aggravated by insistence that the child use speech as do those who hear. Linguistic, cognitive, emotional, and behavioral problems may then follow, because of the attitude and actions of the hearing. All these problems can be resolved by early recognizing that the deaf person has a language and by honestly trying to learn and use it. This language is not a set of signs standing for spoken words but a complete language with its own rules for making sense in signed sentences.


Deaf children can learn to hear.

Deaf children can learn to hear. For more than thirty years the author has been using the "unisensory" approach to work with hearing-impaired children, having learned the principles from her teacher, Emil Froeschels. The approach is based on the theory that even minimal amounts of residual hearing in a hearing-impaired infant, if stimulated in a unisensory fashion, could lead to the development of spontaneous speech and oral communication. Children taught by this method learn to process language through their amplified hearing. They acquire language that is adequate for them to be mainstreamed with their hearing peers from the beginning. The article describes seven requisites to an effective unisensory approach.


A note on simultaneous and interleaved masking. Speech and noise were switched periodically from ear to ear at a rate high enough to create a binaurally fused image. Two masking conditions were investigated. In one the speech and noise were switched in unison from ear to ear such that both speech and noise were presented simultaneoully to the same ear. In the other case, the speech and noise were interleaved such that there was no simultaneous overlap of the speech and noise at any one ear. The difference in masking levels between these two conditions was found to be on the order of 20 dB. The technique offers some interesting possibilities for investigating the relative effects of central and peripheral masking in both normal and hearing-impaired listeners.


Speech and language assessment scales of deaf children. Six year old deaf children with an average PTA of 105.7 dB were evaluated for speech and language skills using rating scales. Five rating scales were designed to assess language production, sign ability, overall communication skills, speech reception and speech intelligibility. Each scale incorporated five categories ranging from inability to complete ability to demonstrate a given skill. For several ratings, classroom teachers, given instruction, were asked to make judgments which were later correlated with examiner's scores, Mean ratings indicate that comprehension skills superceded performance skills. A hierarchy of ability among communication skills can be seen.


Reading attainment in hearing-impaired children: a comparison of higher and lower achievers. The  purpose of the present research is to study hearing-impaired children with higher and lower reading attainment in order to suggest why some of these choldren achieve greater competency than others. Sixteen children, ages 8 to 17, from two oral schools for the hearing-impaired in Britain, were given two tests of reading comprehension. These revealed (1) marked differences in the reading levels of the children, and (2) a relationship between the test scores and what school the children attended. Differences were also found when the children's oral language use and reading comprehension levels were compared. Oral language use was shown to be an important skill related to attainment in reading. Differences in the nature of early language experiences of children in the two schools were said to be partly responsible for the disparity in the children's oral language ability and the differences in their later ability to read. Some insights are offered to explain how one school has fostered the development of good reading skills where other schools have failed to do so.


Pitch deviancy and intelligibility of deaf speech. The relationship between pitch deviancy and other aspects of speech production that affect intelligibiliyy of deaf speech was investigated. The speech skills assessed included the following: ratings of pitch deviancy, ratings of overall intelligibility, production of the prosodic features--stress, intonation, and pause, and production of phonemes. Children who could not sustain phonation had speech that was consistently judged unintelligible. This group of children also had pure tone averages greater than 90 dB. For the remaining children, the relationship among pitch deviancy, intelligibility, and hearing level was variable. The highest intercorrelations were among prosodic feature production, phoneme production, and intelligibility.


The effect of selected prosodic errors on the intelligibility of deaf speech. A trained speech pathologist listened to recordings of 40 deaf children, each reading 15 sentences and recorded instances of four types of prosodic errors: adventitious sounds, excessive phoneme duration, pitch breaks, and pauses. The scores were correlated with previously obtained intelligibility scores. It was found that adventitious sounds, except those obviously operating as transitional sounds to ease the movement from one place of articulation to another, had the greatest negative effect on intelligibility, followed by very long duration and pitch breaks. Medium and long pauses, long duration, prologned closure (i.e., long duration plosives), and adventitious transitional sounds had a lesser negative effect. Short pauses had a small positive effect on intelligibility. The method used to identify the errors was one that can hopefully be evolved into an evaluative instrument for practical use in schools for the deaf.


The evaluation of a model speech training program for deaf children. Systematic speech training procedures were employed with a group of 20 hearing-impaired children ranging in age from 7 to 10 years. Using a primarily auditory-oral approach, the speech training program employed a hierarchical sequence in order to teach the suprasegmental patterns of English to the children. The children's performance on the speech training hierarchy was monitored on a daily basis using a checklist system. Based on their rate of progress in the development of speech skills, the children were divided into three groups post hoc: rapid, slow but steady, inordinately slow. Although the system worked well, on the average, approximately one-third of the children encountered serious difficulites at one or more levels of the hierarchy. Examination of the children's background information and audiological data revealed that those children who encountered difficulty had an extremely limited amount of residual hearing and/or problems other than a hearing handicap.


The relatedness of phonological processes of a hearing-impaired child. This is a study, in the generative framework, of the phonology of a 6-year-old, hard-of-hearing child. The study has two main results: (1) the phonological substitutions and delections of this hearing imparied child are basically the same in kind as those found in the speech of younger normals and (2) the phonological processes of the child's system fit into groups of processes, each group operationalizing some phonetic preference of the child. Implications for a general theory of phonologica acquistion are discussed.


[Projections to the inferior olive of the cat. I. Comparisons of input from the dorsal column nuclei, the lateral cervical nucleus, the spino-olivary pathways, the cerebral cortex and the cerebellum]. The present experiments compare the projections from the dorsal column nuclei (DCN), the lateral cervical nucleus (LCN), the spino-olivary pathways (SO), the motor cortex (Ms1) and the cerebellum (CB) to the inferior olive of the cat. A differential labeling strategy was used for these comparisons. It was found that projections from the contralateral DCN and LCN and the ipsilateral SO overlap extensively with each other in the dorsal accessory division of the inferior olive and the caudal half of the medial accessory olive. Projections from the contralateral motor cortex partially overlap these somato-sensory projections but they also extend into the principal division and into the rostral half of the medial accessory olive where other authors have found input from other "motor" sources such as the red n., the globus pallidus and the caudate n. The contralateral cerebellum projects heavily to most of the inferior olive except for a few regions in the caudal portions of the two accessory nuclei. These results show that there is a partial segregation between somatosensory and motor inputs within the inferior olive and that the cerebellar feedback fibers appear to avoid at least parts of those regions where the somatosensory input is heavy. Some of this segregation, particularly within the medial accessory olive and the principal n., correlates with differences in both the olivo-cerebellar connectivity and the cytoarchitecture of these regions. Although the somatosensory and motor inputs are partially segregated, there are also several regions where these inputs overlap. These regions are: (1) an area in the medial half of the rostral portions of the dorsal accessory olive, and (2) a small oval zone in the middle of the caudal half of the medial accessory olive. This overlap provides an anatomical basis for electrophysiological evidence which demonstrates the existence of cells in some of these regions that respond to activation of both the cerebral cortex and the spinal cord (e.g., Crill and Kennedy, '67).


Projections to the inferior olive of the cat. II. Comparisons of input from the gracile, cuneate and the spinal trigeminal nuclei. The present experiments compared the projections to the inferior olive of the cat from the gracile, cuneate and spinal trigeminal nuclei. A differential labeling strategy was used for these comparisons. It was found that all three somatic sensory nuclei project to portions of all three major divisions of the contralateral inferior olive. The spinal trigeminal n. may also project less densely to the ipsilateral medial accessory olive. Projections to the dorsal accessory nucleus (DAO) and the medially-adjacent ventral lamella of the principal nucleus are roughly somatotopically organized. Although there is considerable overlap between the projection zones, the gracile n. projects predominantly to lateral DAO, the cuneate n. projects predominantly to medial DAO, and the spinal trigeminal nucleus pars caudalis projects predominantly to the most medial portions of DAO and the ventral lamella of principal olive. Projections to the medial accessory olive, on the other hand, are not as highly organized. Instead, they overlap extensively within a small egg-shaped area in the middle of the caudal half of the nucleus. Whereas all portions of the gracile and cuneate nuclei project to the inferior olive, only the pars caudalis of the spinal trigeminal nucleus appears to do so. These results were compared with the three available olivocerebellar maps as well as with the available behavioral and electrophysiological evidence on cerebellar somatotopic organization. This comparison indicated that the inputs to the cerebellum from the three second-order somatosensory nuclei via the inferior olive appear to be generally consistent with cerebellar somatotopic organization. This consistency is apparent not only with respect to the longitudinally-organized, vermal and paravermal differences in the anterior lobe, but also with respect to the transversely-organized specific somatotopy of the intermediate zone of the anterior lobe and the paramedian lobule.


The effects of dark rearing on the development of the visual cortex of the rat. The effects of dark rearing on the development of the visual cortex has been studied in Wistar rats, as have the effects of subsequent light exposure on recovery. Five groups of animals were used: (1) light exposed until 30-40 days post partum (dpp) (2) dark reared until 30-40 ddp (3) dark reared until 80-120 dpp (4) dark reared to 21 dpp, then light exposed until 40 dpp (5) light exposed to 21 dpp and then dark reared until 40 dpp. Golgi-Cox impregnations of layer IV stellate cell dendritic fields were analysed and total neuronal and glial counts were also done within layer IV of the primary visual cortex. Normal visual stellate cell dendritic fields were radially organised, with the highest dendritic density being recorded below the soma. In short term visually deprived animals and in the exposed only for 21 dpp and then reared in light until 40 dpp the radial distribution of dendrites was maintained but the peak density shifted to above the soma. In all other experimental groups this abnormal polarisation was still present but not as marked. Measurement of branching indices suggested that these field changes resulted from increased branching and growth in the superficial domain and not from the reorientation of dendrites. Differential glial counts revealed a significantly higher number of microglia in dark reared animals than in controls. Neuronal numbers were not affected.


In situ injection of kainic acid: a new method for selectively lesioning neural cell bodies while sparing axons of passage. The morphologic sequelae following stereotaxic injection into the rat striatum of kainic acid, a conformationally restricted analogue of glutamate, were examined by means of bright field, histofluorescence and electron microscopic techniques. The neuropathologic response to kainate injection occurs in two distinct phases. First, the intrinsic neurons of the striatum undergo a rapid degeneration during the first 48 hours after injection; this is characterized by the sequential loss of cytoplasmic Nissl-substance (chromatolysis), shrinkage of the perikarya, clumping of the nuclear chromatin and finally disruption of the nuclear membrane. Between one and three weeks after injection, a marked proliferation of astrocytes in the gray matter formerly populated by neurons characterizes the second phase. The region of several neuronal loss in the kainate injected striatum is approximately spherical in shape, and its extent is non-linearly related to the amount of kainate injected. The neuropil of the injected striatum is markedly disrupted due to the death of intrinsic neurons and loss of their processes. Yet, histofluorescence microscopy demonstrates that the dopaminergic axons projecting from the substantia nigra do not degenerate in the kainate injected striatum; electron microscope studies indicate that corticofugal fibers traversing the striatum are also unaffected by kainate. Many presynaptic boutons, presumably of extrinsic origin, are intact up to ten days after injection; osmophilic vestiges of postsynaptic elements remain adherent to these boutons. Numerous phagocytic astrocytes are observed throughout the lesioned area. These morphologic studies provide (direct) evidence that in situ injection of kainic acid in brain causes a selective degeneration of neurons with cell bodies in the area of the injection but spares axons that arise from perikarya outside the region but pass through or terminate in the injected area. Thus, in situ injection of kainic acid is a new technique for making selective brain lesions that will be useful for examining neuronal connectivity.


Visual discrimination following partial telencephalic ablations in nurse sharks (Ginglymostoma cirratum). An instrumental conditioning task was used to examine the role of the nurse shark telencephalon in black-white (BW) and horizontal-vertical stripes (HV) discrimination performance. In the first experiment, subjects initially received either bilateral anterior telencephalic control lesions or bilateral posterior telencephalic lesions aimed at destroying the central telencephalic nuclei (CN), which are known to receive direct input from the thalamic visual area. Postoperatively, the sharks were trained first on BW and then on HV. Those with anterior lesions learned both tasks as rapidly as unoperated subjects. Those with posterior lesions exhibited visual discrimination deficits related to the amount of damage to the CN and its connecting pathways. Severe damage resulted in an inability to learn either task but caused no impairments in motivation or general learning ability. In the second experiment, the sharks were first trained on BW and HV and then operated. Suction ablations were used to remove various portions of the CN. Sharks with 10% or less damage to the CN retained the preoperatively acquired discriminations almost perfectly. Those with 11-50% damage had to be retrained on both tasks. Almost total removal of the CN produced behavioral indications of blindness along with an inability to perform above the chance level on BW despite excellent retention of both discriminations over a 28-day period before surgery. It appears, however, that such sharks can still detect light. These results implicate the central telencephalic nuclei in the control of visually guided behavior in sharks.


The DNA content of Purkinje cells in mammals. Nerve cells have generally been assumed to have a diploid DNA content, typical of non-dividing somatic cells. However several reports have suggested that certain nerve cells types, notably Purkinje cells of the cerebellum, are polyploid. Other studies have contradicted these findings, stating Purkinje cells to be diploid. In this paper we reinvestigate the DNA status of Purkinje cells, in a variety of mammalian species. Cell DNA content is measured on tissue smears by Feulgen microspectrophotometry. Results show that for all species examined by us, Purkinje cells have, without exception, a DNA content comparable to that of somatic cells. A critical appraisal of the techniques used in those studies claiming a tetraploid DNA content for Purkinje cells leads us to believe our findings to be correct.


The role of ependyma in regeneration of the spinal cord in the urodele amphibian tail. The new spinal cord formed during tail regeneration in the newt first develops as a caudal extension of the ependymal tube. Neuroblasts and neuroglia subsequently differentiate from cells of the ependymal tube in a proximal-caudal sweep. Descending axons from the cord rostral to the lesion and from newly differentiating neurons travel in channels which are present prior to the ingrowth of axons. The present study confirms previous observations from our laboratory and presents details of the ultrastructural relations of axons and ependymal processes within the cord. The ependymal cell surface facing channels typically forms numerous digitor sheet-like protuberances which extend into the channel lumen. As axons enter the channels in increasing numbers these protuberances partially subdivide the axons into smaller groupings, even occasionally segregating individual axons. At levels where fibers have not yet entered or have most recently entered the ependymal channesl two specializations appear on the ependymal plasmalemma facing the channels and their axons: coated membranes and hemidesmosome-like structures. At more mature levels, where many fibers have already entered the channels, axons in contact with ependymal processes sometimes show synapse-lide vesicle accumulations with associated membrane densities. Coated membranes and hemidesmosome-like structures are lacking at these levels. Our observations suggest that ependymal processes, in addition to providing substrate and direction for regenerating spinal cord axons, may also furnish or exchange more specific information at the morphologically identifiable specializations described above.


Golgi studies of the neurons in layer II of the dorsal horn of the medulla (trigeminal nucleus caudalis). The translucent band which lies just beneath the spinal V tract at the lower end of the spinal trigeminal nucleus (nucleus caudalis) can be divided into three layers. These three layers are distinguished by textural differences in their neuropil and by the morphology and laminar distribution of the axons and dendrites of their neurons. Layer II contains four different kinds of interneurons. Stalked cells are named after their short, stalk-like branches. Their cell bodies are found in greatest numbers in the outer half of layer II. Their coneshaped dendritic arbors extend medially across layers II and III and sometimes extend into layer IV. Their axons form extensive, canopy-like arborizations in layer I. Stalked cells are considered to be excitatory interneurons receiving input on their dendritic spines from primary axonal endings in the layers II and III glomeruli and transferring it to the dendrites of the layer I projection neurons. Layer II contains three kinds of Golgi type II inteneurons, i.e, neurons whose axons branch repeatedly within the confimes of their dendritic arbors. Islet cells similar to those found in layer III (Gobel), '75a), are found in small clusters in layer II. Their dendrites and axons are largely confined in layer II. The dendrites of the arboreal cell burst, in tree-like fashion, into highly focal dendritic arbors confined largely in layer II while the extensive rostral and caudal dendritic arbors of the II-III border cell lie largely in layers II and III with a few branches extending into layer I. The axons of both of these interneurons arborize in layers II and III with a few collaterals extending into layer I. Islet cells, arboreal cells and II-III border cells are considered to be inhibitory interneurons. They are strategically situated to interrupt transmission between primary axonal endings in layers II and III and the layer I projection neurons by altering the output of the stalked cells.


Organization and development of the brain stem auditory nuclei of the chicken: primary afferent projections. The pattern of primary auditory projections to the brain stem of young chickens was investigated using terminal degeneration methods and orthograde transport of horseradish peroxidase (HRP) or tritiated amino acid. Of particular interest was the question of whether nucleus laminaris (NL) receives primary afferents. A study of silver-stained degeneration pattersn in nucleus magnocellularis (NM) and NL at three intervals following unilateral interruption of the cochlear nerve revealed that by 48 hours after the lesion, degenerating terminals were found only in the ipsilateral nucleus angularis (NA), NM and lagenar projection areas but not in NL. Five- and eight-day survival times, however, also revealed degeneration bilaterally in NL. The appearance of terminal degeneration in NL at the longer survival times is attributed to the previously-reported severe and rapid transneuronal degeneration of neurons in NM following deafferentation and not to the presence of cochlear nerve terminals in NL. Injection of HRP or tritiated proline into the basilar papilla produced patterns of labeling similar to that seen in the 2-day degeneration material; HRP reaction product or autoradiographic label were seen only in the ipsilateral NA and NM and in the ipsilateral projection areas of the macula lagena but not in either NL. The patterns of primary auditory projections revealed by the three methods were quite similar to each other and to that previously reported for the pigeon and confirm the conslucion that the laminar nucleus of chickens does not receive primary afferents.


Properties of the enteric nervous system: limitation of access of intravascular macromolecules to the myenteric plexus and muscularis externa. The possible presence of a blood-myenteric plexus barrier similar to the blood-nerve and blood-brain barriers was investigated. The myenteric plexus was found to be an enclosed tubular structure incompletely surrounded by a sheath of supporting cell processes. Capillaries do not enter the plexus. The capillaries which supply the myenteric layer differ in structure from capillaries of other layers of the gut and are non-fenestrated. Tracers, Evans blue labeled albumin or horseradish peroxidase, readily leak out of fenestrated capillaries, but do not readily escape from myenteric capillaries. These capillaries have impermeable junctions that prevent the passage of tracer between endothelial cells. A slow leakage of macromolecules is probably accounted for by transport through endothelial cells within plasmalemmal vesicles. A backup system of phagocytic cells removes this material and prevents the tracers  leaking from the vasculature from reaching detectable concentrations in the extracellular space. Neither tracer was ever found in the myenteric plexus. Therefore, there is a blood-myenteric plexus barrier to macromolecules that resembles the blood-thymic barrier and may be functionally analogous to the blood-brain barrier.


Light microscopic analysis of the kitten retina: postnatal development in the area centralis. A method was devised for morphological localization of the area centralis, and the timecourse of its formation as a structural entity was established. Postnatal differentiation of the retina proceeds as follows: the irregularly laminated ganglion cell layer of the newborn becomes unilaminar everywhere but in the presumptive area centralis, a difference which is first discernible at five to six days of age; the outer nuclear layer is always of the same thickness in the area centralis, while in the periphery the layer thins with time; the outer nuclear layer is always thinner in the area centralis than in the periphery; inner nuclear layer thickness is invariant early in postnatal life, but in the adult it is thicker in the area centralis than in the near temporal periphery; plexiform layers form by two weeks of age and rach adult thickness thereafter. Retinal ganglion cells were measured and the percent distributions of three ganglion cell size classes (6-10 micron; 11-20 micron; 21-35 micron) were determined for the area centralis and near temporal periphery; Mean ganglion cell size is constant in center and periphery through five weeks age, is adultlike in the periphery soon thereafter and in the center sometime after eight weeks of age. The percent distribution of ganglion cells by size class in center and periphery is not adultlike even at eight weeks of age. The implications of these observations and others are discussed relative to postnatal growth of the eye and placement of the area centralis in the retinal field and optic axis. The involvement of retinal cell proliferation, cell growth, ganglion cell dendrite formation and cell shape changes in the expansion of the retina are also discussed.


Sequence of neuron origin in experimentally distorted brains of chick embryos. The effects of proliferative disorders on the time and sequence of neuron origin in the optic tectum were studied in chick embryos in which "overgrowth" had been experimentally induced, Sixteen normal and fourteen abnormal embryos ranging in age from four through ten days of incubation were each given a single dose of thymidine[3H]. Comparisons were then made between 16-day normal and abnormal chick embryos with respect to the distribution tectum midway between its rostral and caudal boundaries. In the normal and abnormal tecta layers I-IV were generated on days 3-6. Differences occurred in the generation of the remaining layers. In the normal embryos layers V-VII arose on days 5-8 and layers VIII-XV on days 4-7; in the abnormal embryos layers V-VII originated on days 6-8 and layers VIII-XV on days 5-8. In addition to the delay in onset of neuron origin, the abnormal tectum also showed distorted lamination, particularly in layers VIII-XV which constitute the superficial regions of the stratum griseum and fibrosum superficiale. Despite the defect in time and sequence of neuron origin, the rostroventrolateral to caudodorsomedial gradient of maturity characteristic of the normal tecta was not disturbed in the abnormal tecta.