Effect of carbimazole induced hypothyroidism and thyroxine replacement on the growth of the long bones in albino rats of different age groups.

To evaluate the effect of carbimazole induced hypothyroidism and thyroxine replacement, on the growth of long bones of albino rats of different age groups. Experimental albino rats were developed with carbimazole and carbimazole plus thyroxine for a period of six weeks. At the end of the experiment the animals were sacrificed, fixed and processed to demonstrate the bony and cartilaginous parts. The ulna and tibia of both sides were measured for intact bone length & diameter and the data compared. The reduction in length and circumference observed, at the end of experiment, in ulna was 10.89%, & 11.94% and in tibia it was 12.52%, 14.81% in carbimazole treated group respectively, while in carbimazole plus thyroxin treated group the reduction in length & circumference of ulna was 1.37% & 1.88% and in tibia it was 1.86% & 3.08% respectively. They were compared to their age matched controls. The reduction in length and circumference in ulna was 5.58% & 6.25% and 6.42% and 5.88% in tibia respectively among the carbimazole treated animals while in the carbimazole plus thyroxine treated animals the reduction was only 0.63% and 3.12% in ulna and 0.91% and 1.06% in tibia respectively. The results show that hypothyroidism and its replacement therapy affects the endochondral as well as periosteal bone growth and results in reduction in length as well as circumference of long bones.

Impact of selenium, iron, copper and zinc in on/off Parkinson's patients on L-dopa therapy.

We have quantitated CSF and serum levels of Selenium, iron, copper and zinc by Atomic absorption spectrophotometer in 36 patients with parkinson's disease all on L-dopa therapy. Out of these 19 showed on or positive response to L-dopa where as 21 patients showed on and off response. These data were compared with 21 healthy controls. The results showed that serum levels of iron, copper and zinc remained unchanged where as in CSF, significant decrease in zinc was found in both on and on/off PD patients indicating the deficiency of zinc which continues in the worsening clinical condition of off patients. The level of copper remained unchanged in both on and on/off PD patients. Iron and selenium increase in CSF of both patients which is a clear evidence of relationship between increased iron and selenium level in brain which could be correlated with decrease in dopamine levels and oxidative stress in PD Patients.

Polyamines as cancer markers: applicable separation methods.

Spermine, spermidine, putrescine and cadaverine are aliphatic amines widely spread in the human body. Their concentrations together with their acetyl conjugates increase significantly in the biological fluids and the affected tissues of cancer patients. Their concentrations decrease with the improvement in the patient's condition on multiple therapy. Various chromatographic techniques are frequently used in monitoring concentrations of di- and polyamines in cancer. Among these techniques, thin-layer chromatography and liquid chromatography using pre- or postcolumn derivatization, separating on a reversed-phase or an ion-exchange column are the most commonly used. Besides, high-resolution capillary column gas chromatography (GC) is increasingly used over packed column GC, and in recent years, capillary zone electrophoresis has also gained some importance in polyamine determinations. The review examines the prospects and the limitations of polyamines as cancer markers using chromatographic and electrophoretic techniques.

Variation in cerebrospinal fluid levels of neuropeptide Y, cholecystokinin and substance P in patients with neurological disorders.

Neuropeptide Y, cholecystokinin (tetra- and octasulphated peptides) and substance P were measured in lumbar cerebrospinal fluid obtained from patients with various neurologic disorders such as Parkinson's disease, cerebrovascular disorders, multiple sclerosis, tuberculous meningitis and aseptic meningitis. These results are statistically compared with healthy results. The results accumulated showed that the data collected can provide the vital information necessary for designing drug therapy.

The neurochemical markers in cerebrospinal fluid to differentiate between aseptic and tuberculous meningitis.

In this study, the use of neurochemical markers in patients with aseptic and tuberculous meningitis has been investigated. The cerebrospinal fluid levels of amino acids, nitrite (a metabolite of nitric oxide), vitamin B12 and homocysteine were quantitated in both groups of patients. Among the amino acids, aspartic acid and glutamic acid both excitatory amino acid, GABA, glycine and tryptophan were all significantly increased in both patient groups whereas decreased level of taurine and increased level of phenylalanine were only found in patients with tuberculous meningitis. The levels of nitrite and its precursor arginine were significantly higher in patients with tuberculous meningitis whereas unchanged levels were found in patients with aseptic meningitis. A significantly increased homocysteine level and a decreased level of vitamin B12 were found only in patients with tuberculous meningitis whereas unchanged levels were found in patients with aseptic meningitis. This indicates that patients with tuberculous meningitis are particularly prone to vitamin B12 deficiency resulting into increased level of HC, and involvement of free radical showing the importance of these biological markers for promoting the possibility for the design of therapeutic approach.

Cholecystokinin peptides in cerebrospinal fluid: a study in healthy male subjects.

The clinical reliability of measuring cholecystokinin (CCK) peptides in the cerebrospinal fluid (CSF) has not been fully elucidated. Therefore, we have assayed CCK-8S and CCK-4 in CSF obtained from 14 healthy male subjects, lumbar-punctured at the L4-5 level following a strictly standardised procedure. CSF concentrations of free CCK-8S and free CCK-4 were used as dependent variables while age, height, body weight, atmospheric pressure and some other factors served as independent variables. It was shown that the CCK-8S ratio between the second (7-12 ml) and first (0-6 ml) CSF fractions, correlated significantly with the atmosphere pressure at the time of puncture. Neither CCK-8S nor CCK-4 displayed concentration gradients in CSF. The CCK-4 levels, expressed as pmol l-1 in the total amount of CSF were found to be positively correlated with the neuraxis distance in the lying position and negatively with the neuraxis distance in the sitting position. Furthermore, CCK-4, expressed as pmol l-1 per min of tapping-time (pmol l-1 min-1), showed a negative correlation with storage time, presumably mirroring a proteolytic process. CCK-8S and CCK-4 intercorrelated positively independently of whether expressed as pmol l-1 or pmol l-1 min-1. In conclusion, the results of this exploratory study indicate that the neuraxis distance (in the sitting and lying positions) and storage-time have to be accounted for when interpreting data on CSF levels of CCK-4. Attention has to be paid to the potential influence of atmospheric pressure on the concentration ratio of CCK-8S.

Nitric oxide synthase pathway may mediate human natural killer cell cytotoxicity.

The present study provides evidence that the human natural killer (NK) cell effector mechanism causing target cytolysis has a requirement for L-arginine. In a deficient medium (DM) containing only salts, buffer system and glucose, NK cell-mediated cytotoxicity was found to decrease by 70% as compared to that obtained in a complete medium (CM). However, adding L-arginine to such DM could restore the activity of NK cells to the normal level. Many other components of CM, such as serum, glutamine and vitamins did not improve NK cell-mediated killing in DM. When all amino acids except L-arginine were added to DM only a partial recovery of NK cell functional cytolysis was seen. L-arginine enhanced the NK cell activity in a dose-dependent manner. Additionally, the inhibitor of both inducible and constitutive nitric oxide synthase, N-monomethyl-L-arginine (L-NMMA) inhibited NK cytolytic activity in DM supplemented with L-arginine indicating participation of nitric oxide (NO). The results also show that the stimulatory effect of L-arginine on human NK cell-mediated cytotoxicity was accompanied by an increase in NO formation as determined by accumulation of nitrite and citrulline. L-NMMA gave a dose-dependent reduction in NO generation as well. The nitrite and citrulline production dose-dependently correlated with not only the concentration of L-arginine in the cultivation medium, but also the enhanced NK cell-mediated cytolysis. Taken together, these findings could define a L-arginine/NO-linked effector mechanism in human NK cells. Nitrite and citrulline were not formed when NK cell-mediated target cell killing took place in a L-arginine-free DM supplemented with additives. Thus, it appears as if human NK cells may cause target cell killing via both NO-dependent and -independent processes.

Dopamine D1 or D2 antagonists enhance inhibition of consummatory ingestive behavior by CCK-8.

A dopamine D1 (SKF-38393, 1 mg)- or D2 (LY-171555, 0.1 mg)-receptor agonist inhibited intake of an intraorally infused solution of sucrose by male rats, a test of consummatory ingestive behavior. Treatment with a D1 (SCH-39166, 0.1 mg) or D2 (raclopride, 0.6 mg) antagonist reversed inhibition by the respective agonist but enhanced the inhibitory effect of cholecystokinin octapeptide (CCK-8; 1.8 micrograms). It was not possible to demonstrate specific effects of D1 and D2 agonists on intake of pellets, a test that does not discriminate consummatory ingestive behavior from appetitive ingestive behavior, i.e., behavior used to obtain food. The results demonstrate specific involvement of dopamine D1 and D2 receptors in inhibition of consummatory ingestive behavior.

Increased cerebrospinal fluid concentration of nitrite in Parkinson's disease.

The concentration of nitrite, a metabolite of nitric oxide (NO), was increased in the cerebrospinal fluid (CSF) of untreated patients with Parkinson's disease and in patients treated with L-DOPA in comparison with a group of patients without dopaminergic dysfunction. There was no difference in the concentration of L-arginine (ARG), a precursor of NO, between the groups. There was a highly significant, linear relationship between the concentration of nitrite and ARG in the CSF suggesting that the production of NO is dependent on the availability of ARG. The results support the possibility that production of NO is increased in the brain in Parkinson's disease.

Inhibition of serotonin synthesis attenuates inhibition of ingestive behavior by CCK-8.

Ingestive behavior was activated in male rats by intraoral infusion of a 1-M solution of sucrose. Injection of cholecystokinin octapeptide (CCK-8; 1.6 or 5.0 micrograms) inhibited ingestion of the sucrose solution and increased the concentration of 5-hydroxytryptamine (5-HT) in the paraventricular hypothalamic nuclei. The inhibitory effect of the low, but not the high, dose of CCK-8 was attenuated by depleting 5-HT in the brain with p-chlorophenylalanine (PCPA; 100 mg/kg for 3 days). Treatment with 5-hydroxytryptophan (20 mg/kg) increased the concentration of 5-HT in the brain of rats pretreated with either NaCl or PCPA and enhanced the inhibitory effect of CCK-8 on ingestive behavior in the PCPA-, but not NaCl-, treated rats. 5-HT may play a role in the mechanism of action of CCK-8 but additional factors must be involved.

Diet-independent suppression of ingestive behavior by cholecystokinin octapeptide and amino acids.

Male rats consumed much more of an intraorally administered mixed protein, fat and carbohydrate solution than of a carbohydrate solution. Injection of cholecystokinin octapeptide (CCK-8, 0.6-5.0 microgram) suppressed intake of both solutions, but the CCK-A receptor antagonist L-364, 718 (20-40 micrograms) facilitated only carbohydrate intake. Blood levels of CCK-8 were higher after intake of the carbohydrate than the mixed solution. Blood levels of isoleucine, leucine, lysine, threonine, valine, and tryptophan increased only after intake of the mixed solution. Injection of these amino acids suppressed intake of both solutions. Blood levels of amino acids were also less after the seventh than after the first session ingesting the mixed solution. Treatment with CCK-8 or amino acids inhibits intake of any diet, but when secreted endogenously, these signals may terminate the meal in a diet-dependent manner.

Glutamate inhibits ingestive behaviour.

Male rats treated with reserpine were motionless and ingested only a few of ten consecutive intraoral injections of a 1 M solution of sucrose. While injection of apomorphine, a dopamine agonist, stimulated locomotion and stereotyped sniffing in reserpinized rats, it did not reactivate ingestive responses. The non-competitive N-methyl-D-aspartate receptor antagonist MK801, however, stimulated locomotion as well as ingestion suggesting involvement of glutamate in the suppressive effect of resperpine on ingestive responses. A series of experiments was therefore undertaken to investigate the possible physiological role of glutamate in feeding. For this purpose, we used Grill's intraoral intake test, in which the rat is infused intraorally with a sucrose solution and the amount ingested measured. In untreated rats, MK801 dose-dependently facilitated ingestion of the sucrose solution and antagonized inhibition of ingestion by cholecystokinin octapeptide. Administration of cholecystokinin octapeptide or ingestion of sucrose increased the concentration of glutamate in the nucleus of the solitary tract, a brain stem relay transmitting sensory information from the gastrointestinal tract to the forebrain. MK801 was found to bind specifically to this brain area and block the elevation of glutamate and dopamine levels which occurred after treatment with cholecystokinin octapeptide in this neural site. Together these data suggest that dopamine and glutamate may interact within the nucleus of the solitary tract in controlling ingestive behaviour.

Brain cholecystokinin tetrapeptide levels are increased in a rat model of anxiety.

Rats exposed to the smell of a predator adopted the freezing posture indicative of anxiety. Correlatively, the concentration of cholecystokinin tetrapeptide (CCK-4) was increased in the olfactory bulb, frontal and central cortex, dorsal and ventral striatum, central amygdala and the nucleus of the solitary tract. The concentration of CCK-8 was increased only in the ventral striatum. Glutamate was increased in the cortex and the striatum and dopamine was increased in the cortex. Intraperitoneal injection of CCK-4 increased brain levels of CCK-4 and replicated, in part, the behavioural effect of the smell of the predator. Injection of a CCK-B-receptor antagonist had the opposite behavioural effect. The results support a role for CCK-4 in anxiety.

Quantitation and identification of two cholecystokinin peptides, CCK-4 and CCK-8s, in rat brain by HPLC and fast atom bombardment mass spectrometry.

A sensitive HPLC method has been described for quantitation of two cholecystokinin (CCK) peptides in discrete rat brain regions. Separation and quantitation was performed by the reversed-phase HPLC combined with electrochemical detection. Analytical recoveries of the tetrapeptide (CCK-4) and octapeptide-sulphate (CCK-8s) were 96% and 94%, respectively. The between assay coefficient of variation (CV) was less than 3% for both peptides. The within-assay CV was 4% and 6% for CCK-4 and CCK-8s and the detection limit was 2 and 10 pmol/mL, respectively. For identification of structures, the peptides were fractionated by semi-preparative HPLC using a novel SMART system for micropurification. The fractions were analysed by fast atom bombardment mass spectrometry (FAB MS) which confirmed the presence of both CCK-4 and CCK-8s in the rat brain tissue.

The role of tryptophan, 5-hydroxy indole-3-acetic acid and their protein binding in uremic patients.

An on-line high performance liquid chromatography (HPLC) method of analysis with electrochemical detection was developed to quantitate catecholamines, tryptophan (Trp) and 5-hydroxy indoleacetic acid (5-HIAA) in muscle and plasma samples from healthy subjects and uraemic patients undergoing continuous ambulatory peritoneal dialysis (CAPD). Equilibrium dialysis was performed on the plasma samples from 14 healthy and 14 uraemic subjects for the separation of free Trp and its metabolites from bound Trp. The results demonstrate abnormal Trp-albumin binding probably with accumulation of indolic metabolites and abnormally high muscle Trp concentration in uraemic patients.

Application of high performance liquid chromatography in study of sulphur amino acid metabolism in uremic patients.

High-performance liquid chromatography (HPLC) was used for the quantitation of the free amino acids in plasma and muscle samples from 34 patients with chronic renal failure; of these patients, 18 were treated by hemodialysis (HD) and 16 by continuous ambulatory peritoneal dialysis (CAPD). Depletion of taurine was observed in plasma and muscle of uremic patients, whereas methionine was normal. Cysteine sulphinic acid was present in plasma of all uremic patients. The results suggest that taurine depletion is due to decreased endogenous synthesis in uremic patients.

Influence of a meat-free diet on the urinary excretion of 3-methylhistidine and creatinine in chronic renal failure.

Urinary excretion of 3-methylhistidine has been proposed as an index of internal protein breakdown provided that the intake of exogenous 3-methylhistidine (meat) is excluded. To evaluate the potential use of 3-methylhistidine in the metabolic assessment of patients with advanced renal failure, we studied a group of 11 patients with markedly reduced renal function who were put on a meat-free diet (the protein intake was kept constant). Steady-state plasma concentration and urinary excretion of 3-methylhistidine were not achieved until 14 d after exclusion of meat from the diet. At that time the plasma concentration and urinary excretion of 3-methylhistidine had decreased by 43% and 60%, respectively. We conclude that the delay in reaching steady state makes the clinical use of urinary excretion of 3-methylhistidine in patients with advanced renal failure unpracticable as an index of protein breakdown. The exclusion of meat also resulted in a continuous decrease in the plasma level and urinary excretion of creatinine, with the result that plasma creatinine or its reciprocal become misleading for evaluation of changes in renal function until a new steady state has been reached.

A comparison between the effect of cholecystokinin octapeptide and apomorphine on ingestion of intraorally administered sucrose in male rats.

The involvement of dopamine receptors in the inhibitory effect of Cholecystokinin octapeptide on ingestive behaviour was investigated. Male rats were infused intraorally with a 1 M solution of sucrose and the amount ingested after treatment with the dopamine receptor agonist apomorphine was compared with that after treatment with Cholecystokinin octapeptide. The test allows a distinction between the consummatory aspects of ingestive behaviour, i.e. responses used to ingest food, from the appetitive aspects, i.e. responses used to obtain food, because it ignores the latter aspects. Comparisons were also made between the effects of apomorphine and Cholecystokinin octapeptide on pellet intake, a test in which the rat has to display appetitive ingestive behaviour. Injection of apomorphine (400 µg) increased the concentration of plasma apomorphine within 0.3 min and the concentration of dopamine in the cerebrospinal fluid within 1 min of injection and induced behavioural stereotypes within 10 min in food-deprived male rats. Plasma apomorphine and cerebrospinal fluid dopamine levels had decreased by 30 min and the behavioural stereotypies had decreased by 40 min after the injection. Injection of apomorphine also inhibited the consumption of food pellets and the ingestion of sucrose. Inhibition of pellet and sucrose ingestion paralleled the effect of apomorphine on Stereotypie behaviour. Thus, injection of a dopamine receptor agonist is followed by alterations in plasma levels of the agonist, cerebrospinal fluid dopamine levels and in Stereotypie and ingestive behaviour which occur in parallel, in an inverted U-shaped manner and with a temporal delay between each event. These results show a close correlation between dopamine receptor stimulation and inhibition of ingestive behaviour. However, reversal of the inhibitory effect of apomorphine on ingestive behaviour required pretreatment with a lower dose of a dopamine receptor antagonist (cis-flupentixol) (0.1 mg) than reversal of Stereotypie behaviour (0.8 mg). The effect of dopamine receptor stimulation on consummatory ingestive behaviour is thus relatively weak and not secondary to the induction of Stereotypic behaviour. Treatment with a high dose of cis-flupentixol (0.8 mg) caused a prolonged period of immobility but had no effect on the ingestion of sucrose. Dopamine receptor blockade, therefore, interferes with appetitive, but not consummatory ingestive behaviour. Injection of Cholecystokinin octapeptide (5 µg) suppressed pellet and sucrose intake in a manner comparable to that of apomorphine, but induced no behavioural stereotypes and caused a gradual, rather than inverted U-shaped, increase in the concentration of dopamine in the cerebrospinal fluid that did not correlate with the effect on ingestive behaviour. Furthermore, while the inhibitory effect of apomorphine on the ingestion of sucrose was reversed by pretreatment with a low dose of cis-flupentixol (0.1 mg), the inhibitory effect of Cholecystokinin octapeptide was only partially reversed by cis-flupentixol and a higher dose (0.8 mg) was required. Blockade of cholecystokinin-A receptors, by treatment with L-364,718, but not cholecystokinin-B receptors, by treatment with L-365,260, blocked the inhibitory effect of Cholecystokinin octapeptide and, by itself, L-364,718 increased the amount of ingested sucrose. The inhibitory effect of Cholecystokinin octapeptide on consummatory ingestive behaviour, which is mediated by cholecystokinin-A receptors, is likely to involve mechanisms in addition to dopaminergic ones.

Evidence that Release of Dopamine in the Brain is Involved in the Inhibitory Effect of Cholecystokinin Octapeptide on Ingestion of Intraorally Administered Sucrose in Male Rats.

To study the possibility that release of dopamine in the brain mediates the inhibitory effect of Cholecystokinin octapeptide on ingestive behaviour, the effect of amphetamine on intake of pellets or an intraorally administered sucrose solution was compared with that of Cholecystokinin octapeptide. Additionally, comparisons were made between the effect of Cholecystokinin octapeptide and pargyline, a monoamine oxidase inhibitor, and α-methyl-ρ-tyrosine, a tyrosine hydroxylase inhibitor. While amphetamine dose-dependently inhibited pellet intake it failed to inhibit sucrose intake in doses which caused behavioural stereotypies (<800 µg). Cholecystokinin octapeptide (5 µg) inhibited ingestive behaviour in both tests. A very high dose of amphetamine (2 mg) was required to inhibit sucrose intake to a level comparable to that of Cholecystokinin octapeptide. Pargyline (5 to 25 mg) or α-methyl-p-tyrosine (25 to 100 mg) dose-dependently inhibited pellet intake but had only weak effects on the intake of sucrose. Pargyline increased the concentration of dopamine and 3-methoxytyramine in the dorsal striatum and decreased the concentration of 3,4-dihydroxyphenylacetic acid. α-Methyl-ρ-tyrosine decreased the concentration of dopamine and 3,4-dihydroxyphenylacetic acid, but not that of 3-methoxytyramine. Injection of amphetamine (2 mg), but not Cholecystokinin octapeptide, in rats pretreated with pargyline increased the concentration of 3-methoxytyramine in the dorsal striatum and this effect was blocked by pretreatment with α-methyl-ρ-tyrosine. Pretreatment with α-methyl-ρ-tyrosine partially reversed the inhibitory effect of Cholecystokinin octapeptide on sucrose ingestion, enhanced the effect of amphetamine but did not affect that of apomorphine, a dopamine agonist. The results support the possibility that the inhibitory effect of Cholecystokinin octapeptide on consummatory ingestive behaviour, in part, is mediated via release of dopamine in the brain.