Importance of photoperiods in determining temporal pattern of annual testicular events in rose-ringed parakeet (Psittacula krameri).

Male rose-ringed parakeets (Psittacula krameri) were transferred to a long photoperiod (LP; LD 16:8) or a short photoperiod (SP; LD 8:16) for 45 or 90 days on four dates corresponding to the beginnings of different reproductive phases in an annual testicular cycle, and testicular responsiveness was evaluated by comparison with the testicular volume, weight, seminiferous tubular diameter, and germ cell profiles of birds in a natural photoperiod (NP). Exposure of birds to LP during the progressive phase (November) led to precocious maturation of testes after 45 days, but induced regression at 90 days. After showing retarded gametogenic functions at 45 days, parallel (November) SP birds exhibited an accelerated rate of germ cell formation at day 90. During the prebreeding phase (January), there were no remarkable differences in any features of testes among NP. LP, and SP birds at 45 days, but gonadal involution in LP parakeets and active spermatogenesis in SP birds occurred after 90 days. The testes did not show any response to LP or SP for 45 and 90 days when the birds were transferred to altered photoperiods during the breeding (March) and preparatory (June) phases, indicating that the parakeets were photorefractory for at least 6 months (March through September). The results also suggest that initiation and termination of seasonal gametogenic activity in parakeets are possibly functions of endogenous rhythmicity or extraphotoperiodic environmental factors. Duration of light may have certain influences on the attainment of annual peak in spermatogenesis, but in all probability the species has a low photoperiod threshold for induction of testicular growth.

Quantitation of serum tobramycin concentration using high-pressure liquid chromatography.

A high-pressure liquid chromatography method for the quantitative determination of tobramycin in serum is described. The antibiotic was separated from serum by chromatography on a silica gel column. The adsorbed antibiotic was derivatized with o-phthalaldehyde, and then eluted with isopropanol. The derivatized tobramycin was separated by reverse-phase chromatography and quantitated by fluorometry. Serum concentrations as low as 0.5 microgram/ml could be accurately measured. A linear response for serum samples containing tobramycin ranging from 0 to 20 microgram/ml was obtained. Other antibiotics, including various aminoglycosides, did not interfere with the tobramycin assay. Comparison with a standard microbiologic assay gave a correlation coefficient of 0.99. This chemical assay is sensitive, precise, specific, and can be performed in 30 minutes.

Involvement of cyclic guanosine monophosphate (cGMP) and cytosolic guanylate cyclase in the regulation of synaptic ribbon numbers in rat pineal gland.

In the rat pineal gland N-acetyltransferase (NAT) activity and synaptic ribbon (SR) numbers display a circadian rhythm. It is well-known that NAT activity is regulated by adrenergic mechanisms involving cyclic adenosine monophosphate (cAMP) as a second messenger. However, the mechanism involved in the regulation of SR numbers has not been established so far. In the present in vitro study, we have investigated the effects of 8-bromo-cyclic guanosine monophosphate (8-bromo-cGMP), a cyclic guanosine monophosphate (cGMP) analog, and stimulation of guanylate cyclase on SR numbers. Incubation with 8-bromo-cGMP increased SR numbers in a dose- and time-dependent manner. Further, stimulation of the cytosolic guanylate cyclase also resulted in increased SR numbers. Adrenergic agonists stimulated cGMP but did not alter SR numbers. These findings suggest that cGMP is involved as a second messenger in the regulation of SR numbers. Since the adrenergically stimulated increase in cGMP did not influence SR numbers, a non-adrenergic cGMP metabolic pathway seems to be involved in the regulation of SR numbers in the rat pineal gland.

Comparison of human urine gastric inhibitor (HUGI) and bacterial endotoxin as inhibitors of acid secretion.

In conscious cats with gastric fistulas, 10 micrograms . kg-1 of human urinary gastric inhibitor (HUGI) given as an intravenous bolus injection increased mean rectal temperature 1.4 degree C and inhibited mean gastrin-stimulated acid secretion by 64%. The sample of HUGI contained an amount of beta-hydroxymyristic acid corresponding to a 5% contamination of the HUGI with bacterial endotoxin. Injection of bacterial endotoxin in an amount corresponding to the beta-hydroxymyritic acid content of HUGI mimicked, both in magnitude and time course, the increase in body temperature and the inhibition of acid secretion produced by HUGI. We conclude that inhibition of acid secretion by HUGI may be due to the presence of an endotoxin-like contaminant.

Role of light in the mediation of acute effects of a single afternoon melatonin injection on steroidogenic activity of testis in the rat.

Young adult male rats, maintained either in an LD 12 : 12 or in continuous illumination (LL) for one week, were given a single injection of 25 microg melatonin/100 g body wt or ethanolic-saline (control) at 17.00 h. Animals from each group were sacrificed at 11.00 h on the following day. The activity of two important steroidogenic enzymes, 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) and delta(5)-3 beta-hydroxysteroid dehydrogenase (delta(5)-3 beta-HSD), and serum concentrations of testosterone, were measured following highly specific and sensitive spectrophotometric techniques and RIA, respectively. A significant decrease in the activity of both the steroidogenic enzymes was noted in the testes of melatonin-treated rats maintained under normal light-dark schedules, but this response was found to be lacking in the LL rats. However, no significant changes in the level of serum testosterone were noted in either group of melatonin-treated rats from the values in respective groups of ethanolic saline-administered LD and LL rats. Exposure of ethanolic saline-injected rats to continuous light also did not cause any change in the steroidogenic activity of the testis from those in LD rats. The study indicates that continuous light as such does not affect the endocrine function of testis but abolishes suppressive effects of melatonin on the steroidogenic activity of the testis in rat.

Histological, histochemical and biochemical effects of cadmium chloride in female koel (Eudynamys scolopacea).

Effects of single subcutaneous injection (1.5 mg/100 g body weight) of cadmium chloride were studied in the ovary, oviduct, thyroid and adrenal gland of Indian koel (Eudynamys scolopacea). Treatment resulted the significant decrease (p less than 0.001) in the weight of ovary and oviduct. The stromal tissue showed hyperaemia and profused haemorrhage leading to some cellular destruction. A marked degeneration was noticed in the lamina propria of magnum. No remarkable change occured in thyroid histology. An increment of alkaline phosphatase activity was pronounced in ovarian tissue as well as in magnum after the injection. An overall reduction of periodic acid Schiff reaction was noticed in both ovary and oviduct after the salt treatment. The experiment caused a significant reduction of eponephrine content (p less than 0.001) in hypertrophied adrenal medulla. The cortical tissues, however, unaltered in their histomorphology. The loss of total amount of cholesterol (p less than 0.05) from the adrenal gland after the experimentation was recorded. It has been suggested that the augmentation of epinephrine secretion suppressed the gonadal acitivty.

Castration and testosterone induced changes in the pinealocytes of roseringed parakeet, Psittacula krameri, during different phases of the annual testicular cycle.

The pinealocytes in male roseringed parakeets (Psittacula krameri) were studied following bilateral castration and/or therapeutic administration of testosterone during the preparatory (June-July), progressive (Nov.-Dec.), pre-breeding (Jan.-Feb.) and breeding (March-April) phases of the annual testicular cycle. The responses of the pineal to either treatment were found to be almost identical throughout the investigation. In each reproductive phase, the pineal appeared to be hypertrophied following castration and the effect was reversed by therapeutic administration of testosterone, while hormonal treatment to the intact parakeets induced regressive changes in the pinealocytes. Collectively, the results of the current study support the hypothesis that the testis through its hormone testosterone exerts inhibitory influences on the activity of pineal, and may thus be considered as being involved in the determination of an inverse relationship between the pineal and the testis during the annual cycle of free-living parakeets.

Influences of graded dose of melatonin on the levels of blood glucose and adrenal catecholamines in male roseringed parakeets (Psittacula krameri ) under different photoperiods.

Effects of daily evening (just before the onset of darkness in a 24 h light dark cycle) administration of graded doses (25, 50, or 100 microg/100 g body wt./day for 30 days) of melatonin on the concentrations of blood glucose and adrenal catecholamines were studied in sexually active male roseringed parakeets under natural (NP; approximately 12L: 12D) and artificial long (LP; 16L: 8D) and short (SP; 8L: 16D) photoperiods. Blood samples and adrenal glands were collected from each bird during the mid-day on the following day of the last treatment. The concentrations of glucose in blood and epinephrine (E) and norepinephrine (NE) in the adrenals were measured. The results of the study indicated that exogenous melatonin induces hypo- or hyperglycemia depending on the dose of hormone administered as well as to the length of photoperiod to which birds were exposed. The levels of E and NE in the adrenals were shown also to vary in relation to photoperiod and the dose of melatonin administered. But the nature of the influence of melatonin becomes different under altered photoperiodic conditions. It appears that short photoperiods are more effective than long photoperiods as a modulator of glycemic and adrenal catecholaminergic responses to exogenous melatonin. A statistically significant correlation between the levels of blood glucose and that of E and NE in the adrenals was found in the control birds, but not in the melatonin treated birds. The results suggested that the responses of blood glucose and adrenal catecholamines to the treatment with melatonin in the roseringed parakeets may not be dependent on each other.

Diurnal sensitivity of the testes to exogenous testosterone propionate in sexually regressed whitethroated munia Lonchura malabarica.

The testes in different groups of whitethroated munia (Lonchura malabarica) were studied following exogenous treatment of testosterone propionate (1 mg/100 g body wt./day) at 06.00 h or 14.00 h or 22.00 h for 15 consecutive days during the quiescent phase of the annual testicular cycle. The testicular conditions in each group of testosterone treated and appropriate control (administered only with the vehicle of hormone at corresponding time-points) group of birds were analyzed by the studies of paired testicular weight, seminiferous tubular diameter, spermatogenetic index, and cytology of the testicular germ cells. The results of the study revealed that exogenous testosterone was stimulatory to the testicular functions irrespective of the time of treatment. The gametokinetic response of the testes to exogenous testosterone also did not show any statistical difference among the birds treated at different hours of the day. The present study demonstrates for the first time that the responses of the testes to exogenous testosterone during the post-breeding phase in an annual testicular cycle of a wild bird does not vary with the time of administration.

Influence of photoperiods on glycemic and adrenal catecholaminergic responses to melatonin administrations in adult male roseringed parakeets, Psittacula krameri Neumann.

Effects of daily (one hour prior to onset of darkness) injection of melatonin (25 micrograms/100 g body wt. for 30 days) on concentrations of blood glucose and adrenal catecholamines were studied in adult male roseringed parakeets, P. krameri under both natural (NP; about 12L:12D) and artificial long (LP; 16L:8D; lights were available in between 0600 and 2200 hrs) or short (SP; 8L:16D; lights were available between 0600 and 1400 hrs) photoperiodic conditions. The results indicate that neither LP, nor SP as such exerts any significant effect on blood glucose titre of control (vehicle of hormone administered) birds. Treatment with melatonin, however, induced hyperglycemia in both NP and LP bird groups, but hypoglycemia in SP birds. Unlike glycemic levels, amount of epinephrine (E) and norepinephrine (NE) in adrenals of control birds exhibited significant changes under altered photoperiods. A decrease in E and an increase in NE were noted in adrenals of both LP and SP birds. Exogenous melatonin in NP birds also caused a decrease in E and concomittant rise in NE levels. On the other hand, treatment of melatonin in both LP and SP bird groups resulted in an increase in the quantity of both E and NE compared to respective values in adrenals of melatonin injected NP birds. However, relative to the amount of E and NE in adrenals of placebo treated LP and SP birds, significant effect of melatonin treatment was observed only in SP birds. The results suggest that influences of exogenous melatonin on the levels of both blood glucose and adrenal catecholamines are largely modulated by short rather than long photoperiods.

Effects of progesterone on some brain neurotransmitters in intact rats.

Effects of progesterone on four neurotransmitters (viz, noradrenaline, 5-HT, dopamine and histamine) of brain were seen in rats with intact ovaries. It was found that progesterone lowers the noradrenaline concentration in medulla, pons, midbrain, hypothalamus, thalami and pituitary, uniformly, when the rats were killed within 4 hours of progesterone injection. At longer intervals (48 hrs) effects of progesterone were seen when progesterone in heavy dose was administered to rats pretreated with estrogen. It is likely that one of the modes of action of the oral contraceptives may be the reduction of noradrenaline content in selected areas of brain, by progesterone. It is also suggested, therapeutic usage of progesterone carries the risk of development of depression in the user.

Second primary cancers in patients with squamous cell carcinoma of the skin.

The occurrence of second primary cancers was explored in patients with squamous cell cancer of the skin (SCC). The excess incidence subsequent to SCC was mainly in cancers related to sunlight and smoking, and in lymphoproliferative malignancies, it was largest (10-fold) in salivary gland cancer.

Influence of methyl parathion on gametogenic and acetylcholinesterase activity in the testis of whitethroated munia (Lonchura malabarica).

Adult male whitethroated munias, Lonchura malabarica (Aves; Passeriformes), were orally administered with methyl parathion (O, O-dimethyl O-(4-nitrophenyl) phosphorothioate), an extensively used organophosphate pesticide, in graded sublethal dose (5 micro g-, or 10 micro g-, or 20 micro g/100 g body wt/day) for variable durations (1-, 5-, or 10 day/s) during their peak reproductive activities in an annual gonadal cycle. No subtle changes in the feeding behavior, mobility, and body weight were noted between the control and different groups of pesticide-fed birds. As a result of the treatment, the paired testicular weight became reduced significantly only after 10 days at 10 micro g and 20 micro g dose levels, but significant decrease in the number of tubules containing healthy germ cells occurred even after single administration of methyl parathion (MP) at the lowest dose (5 micro g/100 g). With the increase in dose and progress of treatment, the number of tubules with healthy germ cells became gradually decreased. The activity of acetylcholinesterase (AChE) in both the brain and testes of MP-treated birds was inhibited in a dose and duration dependent manner. A significant negative correlation was observed between the number of tubules containing degenerated germ cells in the testis and the AChE activity in both the brain and testes of MP-administered birds. However, no remarkable changes in the cytomorphological features, including the nuclear diameter of Leydig cells, were noted in any testis of the pesticide-treated munias. The results of the present investigation suggest that methyl parathion ingestion is harmful to male gametogenic functions in the studied passeriform bird, and the given pesticide may exert its antigonadal effect by impairing cholinergic functions of the brain and/or testes.

Development of day-night rhythmicity in "synaptic" ribbon numbers in the pinealocytes of posthatch chicks kept under either natural photoperiodic conditions or continuous illumination.

Pineal synaptic ribbons (SR) undergo characteristic changes over a period of 24 hr under natural photoperiodic conditions in various vertebrates, being low in number during daytime and elevated at night. During posthatch development of chicks, the rhythmicity of SR numbers is reported to appear at the age of about 2 weeks. Because the influence of external light during the growth phase of chicks on the development of day-night rhythmicity in SR numbers is unknown, we studied day-night differences in SR numbers in the pinealocytes of chicks at the posthatch ages of 15, 17, and 19 days; chicks had previously been kept under natural photoperiodic conditions or continuous illumination. Under natural photoperiodic conditions a statistically significant nocturnal (midnight) rise in SR numbers over the value of midday was seen in the pineal of 17- and 19-day-old chicks, but not in 15-day-old chicks. SR numbers in the pinealocytes of continuously illuminated chicks did not show any day-night rhythmicity on days either 15 or 17, but exhibited significant day-night differences on day 19 posthatch. These findings suggest that continuous illumination, which is known to dampen circadian rhythmicity of melatonin secretion in the chick pineal, causes a delay, but not a total suppression of the mechanism involved in the ontogenic development of diurnal rhythmicity in SR numbers in the pinealocytes of chicks.

Heterogeneity and refined structtures of 3-O-methyl-D-mannose polysaccharides from Mycobacterium smegmatis.

The 3-O-methyl-D-mannose-containing polysaccharide (MMP) from Mycobacterium smegmatis, first described by Gray and Ballou (Gray, G. R., and Ballou, C. E. (1971) J. Biol. Chem. 246, 6835-6842) is now shown to be a mixture of at least four isomers separable by gel filtration owing to differences in size and degree of methylation. The major component is 3-O-methylmannose but all contain small amounts of mannose. The molecular weights range from 2040 to 2490 and all are nonreducing. After Smith degradation, all yield a single large and one or more small fragments that give 3-O-methylmannose as the sole product of complete acid hydrolysis. The large Smith-degraded MMP components (SD-MMP) are similar to intact MMP and vary from 1830 to 2130 daltons, consistent with the loss of a single mannose; whereas the smaller fragments are the size of tri- to hexasaccharides and result from fragmentation of incompletely methylated chains. Controlled acid hydrolysis of [methyl-3H]MMP releases 6% of the methyl groups as [3H]methanol at a rate characteristic for the hydrolysis of methyl alpha-D-mannopyranoside. Proton magnetic resonance spectra of MMP and SD-MMP show a major methyl ether proton peak and a second small peak at higher field equivalent to about one methyl group per molecule. The results are consistent with the presence of an alpha-methyl aglycon at the reducing end of the chains. Methylation analysis of MMP isomers purified by high pressure liquid chromatography confirms that they are linear and unbranched. Methylation of [methyl-3H]MMP yields unlabeled tetra-O-methylmannose, showing that the chains are terminated by mannose. However, digestion of [methyl-3H]MMP with alpha-mannosidase releases mannose and exposes [methyl-3H]3-O-methylmannose. Smith degradation of [methyl-3H]MMP III yields a penta-to hexasaccharide product that can be resolved by high pressure liquid chromatography into two components. The distribution of radioactivity between these two fragments suggests that the chain was cleaved near the middle and that there must be an unmethylated mannose at that position. We conclude that the 3-O-methylmannose polysaccharides are linear unbranched chains of 11 to 14 sugar units, each terminated by a single mannose at the nonreducing end and by a methyl aglycon at the reducing end. Each isomer shows microheterogeneity, with 1 or 2 unmethylated mannose units near the middle of some but not all of the chains.

Characterization of a mannan-like oligosaccharide from Mycobacterium smegmatis.

A nitrogen-free neutral mannooligosaccharide, similar in structure to the polysaccharide component of yeast mannoproteins, has been isolated from Mycobacterium smegmatis ATCC-356. It has a molecular weight of 3200 and is terminated at the reducing end by mannose. nuclear magentic resonance spectroscopy, methylation analysis, selective enzymic degradation and acetolysis indicates that the molecule consists of an alpha1 --> 6-linked backbone to which single mannose units are attached in alpha1 --> 2 linkage as sidechains.

Establishment of beta-hydroxy fatty acids as chemical marker molecules for bacterial endotoxin by gas chromatography-mass spectrometry.

Selected ion-monitoring gas chromatography-mass spectrometry was used for detection of beta-hydroxy fatty acids as an independent assay for the presence or absence of endotoxin in materials claimed to induce nonspecific activation of Limulus amoebocyte lysate. To this end, suspensions of gram-negative and -positive bacteria, one fungal species, cerebrospinal fluid, and hollow-fiber hemodialyzer rinses were assayed for endotoxin by gas chromatography-mass spectrometry and the Limulus amoebocyte lysate assay. Good qualitative agreement was shown for both methods when suspensions of test organisms were assayed. Two false-negative results were obtained by gas chromatography-mass spectrometry assays of cerebrospinal fluid and were shown to be a result of insufficient endotoxin in the cerebrospinal fluid specimens for detection by gas chromatography-mass spectrometry. Hemodialyzer rinses were Limulus assay positive; however, no beta-hydroxy fatty acids were detected by gas chromatography-mass spectrometry. These data were compared with data obtained from USP rabbit pyrogen tests of the rinse materials (nonpyrogenic) and chemical characterization of the Limulus assay-reactive rinses, which showed the rinses to be cellulosic in nature. It is suggested that beta-hydroxy fatty acids, as assayed by selected ion-monitoring gas chromatography-mass spectrometry, be used as chemical marker molecules for the presence or absence of endotoxin in materials reported to cause nonspecific activation of Limulus amoebocyte lysate.

The effects of short pulses of light at night on numbers of pineal "synaptic" ribbons and serotonin N-acetyltransferase activity in male Sprague-Dawley rats.

To characterize further the functionally enigmatic "synaptic" ribbons (SR) of the mammalian pineal gland and to study possible relationships to melatonin synthesis, in the present investigation rats were exposed to short pulses of light at night when both SR numbers and serotonin N-acetyltransferase (NAT) activity are high in comparison to day-time values. Male Sprague-Dawley rats were killed at 13:00 and 01:00 h, respectively, and at 01:10 and 02:00 h after exposure to light for 10 and 60 min, respectively. The pineals were rapidly taken out and cut sagittally in half. One half was processed for electron-microscopic quantitation of SR numbers and the other half for NAT determinations. It was found that both SR numbers and NAT activity decreased significantly when the animals were exposed to light at night. Although both parameters showed corresponding changes, there was no clear-cut correlation between SR numbers and NAT activity in individual animals within a group, except after exposure to light for 60 min when a positive correlation (R = 0.939; p less than 0.05) existed. After exposure to light the electron-lucent vesicles of the SR decreased in number, but the length of the SR was unchanged. These results show that numbers of pineal SR can be easily and quickly manipulated and that the presently used model may be ideal in studying the poorly understood mode in which degradation of SR occurs.