The growth of Nicotiana silvestris in suspension culture is inhibited by all of the common protein amino acids at the millimolar level, except for L‐glutamine. A defined experimental system for growth/inhibition studies has been established, and growth studies were carried out with cells that had been maintained in the exponential growth phase for at least 10 generations (EE cells). The following results were obtained after particularly detailed studies with aromatic amino acids. The onset of inhibition was preceded by a duration of normal growth rate which varied within a range of 12 to 48 h. The degree of inhibition was directly proportional to amino acid concentration and inversely related to the initial cell density of the inoculum. A slowed, but still exponential rate of growth persisted during an early phase of inhibition. Under sufficiently severe conditions, this was followed by progressive diminution of growth rate and eventual lysis. The most drastic inhibitory effects caused by aromatic amino acids were in the order: phenylalanine, tryptophan and tyrosine. When EE cells cultivated under conditions of growth inhibition were diluted into fresh medium, immediate resumption of growth at the uninhibited rate occurred and persisted. On the other hand, when growth‐inhibited EE cells were diluted into medium containing the same concentration of amino acid used in the first round of growth, an initial burst of uninhibited growth lasting about 24 h was followed by a drastic, progressively declining growth rate which deteriorated to cell death and lysis. When cells in stationary phase were used as an inoculum, as is done in typical growth characterizations with suspension cultures, the sensitivity to inhibition during the subsequent exponential growth phase was several‐fold greater than was the case with EE cells. Hypotheses that growth inhibition might be caused by ammonia toxicity, keto‐acid toxicity, or by inhibition of nitrate utilization were ruled out. Observations that provide new insight are: (i)growth‐inhibited cells undergo drastic plasmolysis, (ii) L‐glutamine is an effective antagonist of amino‐acid inhibitors, and (iii) growth‐inhibited cells exhibit a transient restoration of normal growth rate upon dilution into fresh growth medium. These results implicate a linkage of amino acids with osmotic regulation and nitrogen metabolism.

1992. Amino acids are general growth inhibitors of Nicotianasilvestris in tissue culture. -Physiol. Plant. 84: 319-328.Tbe growth of Nicotiana silvestris in suspension culture is inhibited by all of the common protein amino acids af Ihe millimolar level, except for t-glutamine. A defined experimental system for growth/inhibition studies has been established, and growth studies were carried out with cells Ihat had been maintained in the exponential growth phase for at least 10 generations (EE eells). The following results were obtained after particularly detailed studies with aromatic .amino acids. The onset of inhibition was preceded by a duration of normal growth rate which varied within a range of 12 to 48 h. The degree of inhibition was directly proportional to amino acid concentration and inversely related to the initial cell density of the inoculum. A slowed, but still exponential rate of growth persisted during an early phase of inhibition. Under sufficiently severe conditions, this was followed by progressive diminution of growth rale and eventual lysis. The most drastic inhibitory effects caused by aromatic amino acids were in the order: phenylalanine, tryptophan and tyrosine. When EE cells cultivated under conditions of growth inhibition were diluted into fresh medium, immediate resumption of growth at the uninhibited rate occurred and persisted. On fhe other hand, when growth-inhibited EE cells were diluted into medium containing the same concentration of amino acid used in the first round of growth, an initial burst of uninhibited growth lasting about 24 h was followed by a drastic, progressively declining growth rate which deteriorated to cell death and lysis. When cells in stationary phase were used as an inoculum, as is done io typical growth characterizations with suspension cultures, the sensitivity to inhibition during the subsequent exponential growth phase was several-fold greater than was the ease with EE cells. Hypotheses that growth inhibition might be caused by ammonia toxicity, keto-acid toxicity, or by inhibition of nitrate utilization were ruled out. Observations that provide new insight are: (i) growth-inhibited cells undergo drastic plasmolysis, (ii) L-glutamine is an effective antagonist of amino-acid inhibitors, and (iii) growthinhibited cells exhibit a transient restoration of normal growth rale upon dilution into fresh growth medium. These results implicate a linkage of amino acids with osmotic regulation and nitrogen metabolism.

SUMMARY Prostaglandin release from microvessels isolated from the rabbit cerebral cortex was determined under three different atmospheric conditions: 100% O 2 ("O 2 ") room air, and 95% N 2 =5% CO 2 ("N 2 -COj"). Initial studies with homogenates prepared from rabbit cerebral microvessels (RCMV) indicated two pathways of enzymatic PGH 2 transformation, namely PGI 2 synthase and GSH-dependent PGH-PGE isomerase. We measured the release of the principal products of these pathways, 6-keto PGF |a and PGE, from freshly prepared RCMV. The release of 6-keto PGF |a exceeded that of PGE^ in all three protocols. RCMV incubated in "N 2 -CO 2 " exhibited a reduction in the release of 6-keto PGF la compared to room air or "O 2 " incubated RCMV, evident at 30-60 min of incubation. No significant differences in the release of PGE 2 were observed among the three incubation protocols. In all three incubation protocols the ratio of 6-keto PGF |a to PGE 2 did not differ during the initial 10 minutes of each incubation. After 30 to 60 min of incubation, this ratio did not change from the "O 2 " or room air treated RCMV, but decreased significantly for the "N 2 -CO 2 " treated group. To determine the reversibility of the apparent "N 2 -CO 2 " induced decline in 6-keto PGF |a release, microvessels were removed from the nitrogen atmosphere and incubated in room air. Release was measured during the initial 10 min following reintroduction to room air and was compared to room air pretreated controls treated in an identical manner. Complete recovery of 6-keto PGF |a production was observed, and an enhanced ratio of 6-keto PGF lo to PGE 2 observed in the "N 2 -CO 2 " treated RCMV. In view of the opposing actions of PGI 2 (vasodilator) and PGE 2 (mild vasoconstrictor) on cerebrovascular tone, this study suggests that release of prostaglandins by the microvasculature may participate in the cerebrovascular response to ischemia. In addition, this study suggests that the release of 6-keto PGF |n and PGE 2 by cerebral microvessels may be regulated independently.Stroke Kenzie 9 have shown that injection of indomethacin in baboon induces a decrease in CBF, proposed by these authors to be due to inhibition of prostacyclin synthesis by vascular endothelium. Pickard and Mackenzie 9 also demonstrated a reduction in the CBF response under hypercapnic conditions when indomethacin is administered. However, a recent study by Wennmalm and co-workers 10 indicates the effects of indomethacin on basal and hypercapnic CBF (in man) is not due to the effects of indomethacin on PGI 2 (or other prostaglandins) synthesis and release in the cerebral circulation. In this study, these authors compared the effects of indomethacin, naproxen and aspirin on CBF under basal and hypercapnic conditions. Administration of all three drugs eliminated arachidonic acid induced platelet aggregation in platelets obtained from the treated patients. Only indomethacin, and only when given acutely, elicited a decrease in CBF under basal or hypercapnic conditions. Additionally, these auth...