Individual self-administration of nicotine by rats.

Self-administration (SA) of nicotine (N) was studied in 20 male and 19 female N:NIH rats using the two-bottle method. The experimental protocol consisted of seven consecutive periods each lasting 6 days: Period (P)1, choice of water (W) and 0.003% N; P2, choice of W and 0.006% N; P3, choice of W and 0.012% N; P4, W only; P5, choice of W and 0.006% N; P6, 0.006% N only; and P7, choice of W and 0.006% N. Group means showed that males and female rats consumed similar amounts of N during Ps 1-3. After an N-free period (P4), a small decline was observed in the subsequent voluntary intake of N (P5). Forced N (P6) exposure did not affect a subsequent N intake (P7) in males but increased it slightly in females. A survey of individual animals, however, showed that the voluntary N consumption varied greatly among animals, but was quite consistent for a particular rat. Values ranged from 0.43 to 7.59 for males and from 0.35 to 4.69 mg/kg/day for females for Ps 1-3. The N-free (P4) and the forced-N (P6) periods each affected a subsequent voluntary N intake (P5, P7) of the rats very differently, but again consistently, in that some rats decreased, some increased and some did not change their N choice. The results indicate that group means can be misleading in their conclusions and strongly support the assumption that the response of an individual animal to N, and not N per se, is the determining force of its SA.

Voluntary self-administration of both morphine and cocaine by rats.

Voluntary self-administration of cocaine and/or morphine was studied in rats. Male rats were offered water bottles or bottles containing either cocaine or morphine, both cocaine and morphine (combination) or cocaine and morphine as a mixture. Alternating the three drug-containing bottles had no effect on drug choice. When offered alone, rats consumed about 12 +/- 8 mg/kg/day of cocaine or 0.3 +/- 0.3 mg/kg/day of morphine. When both drugs were offered in combination, they consumed a higher amount of cocaine (22 +/- 7), but the same amount of morphine (0.4 +/- 0.3). Availability of cocaine/morphine mixture kept morphine consumption constant (0.3 +/- 0.1), but markedly decreased cocaine intake (0.3 +/- 0.2). Addition of saccharin to the drug solutions only slightly increased consumption of both drugs, whereas saccharin added as a competitor or distracter to the drug solution reduced cocaine but not morphine self-administration. Animals showed wide interindividual variations but surprisingly small intraindividual variations in self-administration of cocaine or morphine under all conditions. No correlation between cocaine and morphine intake was apparent in the combination situation. Forcing animals first with cocaine had no effect on subsequent intake of cocaine or morphine presented in combination. However, forcing animals first with morphine subsequently increased morphine and reduced cocaine intake. In conclusion, morphine intake was the same if offered alone, in combination or as a mixture, whereas cocaine intake increased during a combination but decreased in the mixture situation. Cocaine pre-exposure had no effect on subsequent voluntary morphine or cocaine choice, whereas morphine pre-exposure increased subsequent voluntary morphine but decreased cocaine intake. These results suggest the possibility of two reward centers, one for each drug, the morphine center exerting a dominant influence over the cocaine center.

Effects of strain, behavior and age on the self-administration of ethanol, nicotine, cocaine and morphine by two rat strains.

Two genetically different strains, Brown Norway rats (BNR) and Wistar Kyoto rats (WKR), with the latter showing higher emotionality and lower plasma stress catecholamine responses, were compared for their voluntary intake of ethanol, nicotine, cocaine and morphine. Younger BNR self-administered the same amounts of all 4 substances as did the younger WKR suggesting a similar genetic basis for all drugs at this age. Older BNR consumed less ethanol and nicotine but equal amounts of cocaine and morphine as compared to older WKR, and older BNR were more sensitive to the effects of ethanol than WKR suggesting a different genetic basis for different drugs at an older age. Forcing both strains to consume one of the drugs did not affect a subsequent voluntary consumption of ethanol and morphine but reduced nicotine intake in WKR and decreased cocaine intake in both strains suggesting that drug use is determined by individual preferences and not drug exposure per se. The behavioral characteristics of both strains coincide only with the self-administration of ethanol and nicotine supporting a possible genetic linkage between anxiety/stress and ethanol and nicotine use.

Oral self-administration of ethanol and cocaine in rats.

Most laboratory animal studies on self-administration of drugs of abuse use only one drug, whereas humans frequently engage in polydrug use. For this reason, we studied oral self-administration of ethanol (E) and cocaine (C) with the free choice bottle method using a single drug alone, a combination (E and C in separate bottles) or a mixture of both drugs in a single bottle. Young female rats (45 days) consumed similar amounts of C if offered alone (12.4 +/- 7.5 mg/kg/day), in the presence of ethanol (10.6 +/- 3.5) or as E/C mixture (8.0 +/- 4.0). They also consumed similar amounts of E if offered alone (3.8 +/- 1.6 ml/kg/day), in the presence of C (2.3 +/- 0.8) or E/C mixture (2.4 +/- 1.1). Voluntary consumption of both drugs varied markedly among animals but was consistent in a given rat. No correlation occurred between consumption of E and C. Young male rats behaved similarly and consumed similar amounts of E and C alone, in combination and as mixture. While E consumption was similar, C consumption was higher in female rats. Old male rats (180 days) were similar to young male rats. The presence of a saccharin solution as a distracter had no effect on intake of E or C in young females but reduced E intake only in young male rats. In young animals, prior voluntary consumption of either E or C had no effect on subsequent voluntary consumption of the same or other drug offered in combination. These results indicate that this model may be useful to study polydrug use in humans, that consumption of both E and C is strongly controlled by an individual animal, that prior exposure to one drug had no or little effect on a subsequent consumption of the same or other drug in combination and that intake of E or C seems to be independent of each other suggesting two independent reward centers.

Cocaine intake by rats correlates with cocaine-induced dopamine changes in the nucleus accumbens shell.

Extracellular dopamine levels were determined by microdialysis in the core and shell of the nucleus accumbens and the frontal cortex of rats before and after an injection of cocaine (20 mg/kg, IP). After removal of the probes, these same animals were then tested for their voluntary intake of cocaine using the two-bottle, free-choice paradigm. Baseline dopamine levels and their responses to an injection of cocaine differed among the three brain areas. No significant correlations were found between baseline dopamine levels in any of the three brain regions and the voluntary cocaine consumption. A significant negative correlation was found between cocaine-induced increases in extracellular dopamine in the shell of the nucleus accumbens and the voluntary intake of cocaine (r = -0.73, p < 0.01). No such correlations were observed in the accumbens core region or the frontal cortex. These results provide further evidence of the role of the accumbal shell region in cocaine preference, and indicate that cocaine-induced increases in dopamine levels play a role in oral cocaine self-administration or preference. In addition, this relatively novel approach in using the same animals for both cocaine induced neurotransmitter responses and cocaine preference studies can also be applied for the study of other neurotransmitters and drugs of abuse.

Determinants of the voluntary consumption of nicotine by rats.

The 2-bottle free-choice method was used to study the voluntary consumption of nicotine by rats. Rats consumed nicotine voluntarily at different, albeit quite consistent, amounts. Voluntary intakes were higher in younger than older rats, but were not affected by gender. A previously forced nicotine intake had no effect on a subsequent voluntary intake of nicotine in older but increased it in younger rats. Forced exposure to nicotine of pregnant and lactating rats did not increase the voluntary intake by their offsprings. Established free-choice drinking patterns of nicotine were not affected by a temporary forced intake of this substance. The 2-bottle choice proved to be a reliable method to study the voluntary intake of nicotine, and results suggest that nicotine is not 'addictive' per se, but that its use is apparently determined by the response of an individual rat to this substance.

Influence of various drugs on the voluntary intake of nicotine by rats.

The effects of the dopaminergic agonists (L-dopa, pergolide) and antagonists (haloperidol, clozapin) and a cholinergic agonist (tacrine) and antagonist (mecamylamine) on the voluntary intake of nicotine were investigated with the 2-bottle paradigm with the test drugs being dissolved directly in the drinking fluid of the animals. This method was found to be a reliable procedure to quickly screen compounds with specific sites of action in the brain for their effects on the voluntary intake of nicotine or perhaps other substances of abuse as well. L-dopa, pergolide and haloperidol did not affect the intake of nicotine, whereas tacrine increased it slightly and clozapine and mecamylamine markedly. These results indicate that blockade of nicotinic and dopaminergic D4 receptors partially reduce the desired effect of nicotine by forcing the animals to consume more of this substance.

Voluntary consumption of amphetamine, cocaine, ethanol and morphine by rats as influenced by a preceding period of forced drug intake and clozapine.

Forced nicotine intake was previously found to decrease a subsequent free choice selection, whereas clozapine (CL) caused a marked increase in its consumption. Here these findings are extended to ethanol, cocaine, morphine and amphetamine. Forced intake of ethanol, cocaine, morphine and amphetamine had no major effect on a subsequent voluntary intake. CL, a dopamine D4 antagonist, increased the voluntary consumption of amphetamine and morphine with no effects on ethanol or cocaine intake. Only for cocaine was it found that low-consuming rats increased but high-consuming rats decreased their voluntary cocaine intake by CL. Thus, forced drug exposure per se does not lead to subsequent enhancement of voluntary intake; CL exerts differential effects on intake of these drugs, and a specific dopaminergic set point may govern the voluntary intake of cocaine by individual rats.

Effects of stress on amino acids and related compounds in various tissues of fasted rats.

The purpose of this study was to examine the effect of stress on the free amino acid pattern of plasma and various organs. Two groups of rats were deprived of food, for 24 hrs. One group was sacrificed after this time (fasting control representing mostly free endogenous amino acids) and the second group was first restrained in wire cages for 120 min before being sacrificed (fasting stress representing mostly the effects of stress on endogenous free amino acids). A third group had free access to food and was sacrificed at the same time (fed control representing mostly free amino acids absorbed from the gut and endogenous free amino acid metabolism). Fasting (as compared to fed controls) reduced alanine and arginine but increased ethanolamine, glutamic acid and glutamine in the plasma; increased ethanolamine, phosphoethanolamine and glutamic acid in the liver; increased carnosine, glutamic acid, phosphoethanolamine and glutamine in the ventricle; increased oxidized glutathione in the aorta; decreased alanine, aspartic acid, glutamic acid, leucine and methionine and increased glutamine in the pancreas; and decreased arginine in skeletal muscle. Fasting plus stress (as compared to fasting controls) reduced alanine and glutamine in the plasma; increased methionine in the liver; increased ethanolamine, GABA, and glutamic acid in the aorta; reduced arginine, glutamic acid, glutamine, leucine and methionine but increased ethanolamine in the ventricle; reduced ammonia and ethanolamine but increased histidine, isoleucine, leucine, lysine, phenylalanine, tyrosine and valine in the pancreas; and reduced ammonia in skeletal muscle. Fasting plus stress affects the amino acid composition of plasma and various of tissues but effects seen were individually different and strongly substance and tissue specific. Plasma changes did not coincide with tissue changes. Changes in the endogenous pattern of amino acids and related compounds in response to stress could be first indications of stress induced organ pathology.

Effects of L-DOPA/carbidopa administration on the levels of L-DOPA, other amino acids and related compounds in the plasma, brain and heart of the rat.

Rats were treated intraperitoneally with a mixture of 250 mg/kg L-DOPA and 40 mg/kg carbidopa or with vehicle and sacrificed 30 min later. Plasma, heart and cortex, midbrain, brainstem and cerebellum were removed from each animal and assayed by HPLC for L-DOPA and a large number of amino acids and related amino compounds. L-DOPA levels increased from undetectable (<0.2 nmol/ml or g) to 1,146, 1,007, 399, 376, 368 and 850 nmol/ml or g in the above tissues. In addition, several amino compounds were significantly affected by L-DOPA/carbidopa (p < or = 0.01). Plasma concentrations of phosphoserine, oxidized glutathione, citrulline, phenylalanine, tyrosine and 1-methylhistidine increased and arginine, glutamic acid and lysine decreased. In the heart, concentrations of phosphoserine, taurine, reduced glutathione, threonine, serine, glutamine, glycine, alanine, valine, GABA, ethanolamine, ammonia and arginine decreased. In the cortex, camosine and homocarnosine increased. In the midbrain, valine increased and leucine, ornithine and oxidized glutathione decreased. In the cerebellum, citrulline increased. In the brainstem, threonine, serine, asparagine, glutamine, oxidized glutathione, alanine, and leucine decreased. In the brainstem, arginine was slightly decreased with a concomitant increase in citrulline (p < 0.05), indicative of nitrous oxide formation. These results show that administration of L-DOPA/ carbidopa not only raises dopamine levels but can also affect other biochemicals and that the observed changes in amino acids and related compounds can perhaps contribute to the beneficial and/or adverse effects of L-DOPA/carbidopa therapy of Parkinson's disease.

Serotonergic receptors modify the voluntary intake of alcohol and morphine but not of cocaine and nicotine by rats.

Effects of fluvoxamine, a relatively selective 5-HT uptake inhibitor, and ipsapirone, a relatively selective 5-HT1A agonist, were studied on the initiation and/or maintenance of the voluntary intake of alcohol, morphine, cocaine, and/or nicotine in rats using the two-bottle free-choice method. Fluvoxamine (30 mg/kg/day in the drinking fluid) when given during existing morphine consumption increased the intake of this drug (1 +/- 1 vs. 3 +/- 1 mg/kg/day) but had no effect on alcohol (2 +/- 2 vs. 2 +/- 2 g/kg/day) or cocaine (10 +/- 10 vs. 13 +/- 10 mg/kg/day) intake. Ipsapirone (10 mg/kg/day in the drinking fluid) when given during existing alcohol or morphine consumption decreased the intake of the first (2 +/- 2 vs. 1 +/- 1 g/kg/day) and increased the intake of the second drug (2 +/- 1 vs. 4 +/- 1 mg/kg/day), but had no effect on nicotine (1 +/- 1 vs. 1 +/- 1 mg/kg/day) or cocaine (7 +/- 8 vs. 7 +/- 6 mg/kg/day) intake. Ipsapirone when given before exposure to the above drugs reduced subsequent alcohol (2 +/- 1 vs. 1 +/- 1 g/kg/day) and increased subsequent morphine intake (2 +/- 2 vs. 4 +/- 1 mg/kg/day), but had no effect on the voluntary consumption of cocaine (8 +/- 7 vs. 10 +/- 6 mg/kg/day) and nicotine (1 +/- 1 vs. 1 +/- 1 mg/kg/day). These results suggest: (1) selective stimulation of 5-HT1A receptors reduces alcohol preference, (2) stimulation of all 5-HT receptors has no effect on alcohol intake, indicating the presence of inhibitory receptors, (3) stimulation of the serotonergic system in general stimulates morphine preference, (4) the serotonin system does not affect nicotine or cocaine preference and (5) the serotonergic system is not involved in the voluntary consumption of all, but-only of some drugs/chemicals of abuse. Recognition of these drug/chemical-specific sites in the brain might lead to a better understanding of differences in drug abuse patterns among humans and help in the development of specific drugs for the treatment of selective drug addictions.

Levels of amino acids and related compounds in bronchoalveolar lavage fluids of asthmatic patients.

The constituents of bronchoalveolar lavage (BAL) fluid have been shown to reflect the presence and possible etiology of several pulmonary diseases. Presently, although research studies have reported the concentrations of cytokines and compounds such as major basic protein in BAL fluids, only the cellular elements, total protein, albumin, and immunoglobulins have been well defined. We hypothesize that amino acids and related amino compounds, well known participants in physiologic and biochemical processes, are present in BAL fluid and may have involvement in asthma. Our objective was to extend knowledge of the total chemical profile and clinical value of BAL fluids in humans by measuring these amino compounds in normal control subjects and asthmatic patients. Analysis by high-pressure liquid chromatography revealed the presence of 25 compounds. A few compounds in control subjects and patients were found to have values > 1.0 nmol/ml, while the majority were present in comparatively low concentrations < 1.0 nmol/ml. Asparagine, phosphoethanolamine, and taurine were significantly increased in the asthmatic patients. We conclude that the present profile of amino acids and related amino compounds in BAL fluid serves as a potential diagnostic tool in the study of various pulmonary disorders. The significance of increased asparagine, phosphoethanolamine, and taurine in the asthmatic patients is discussed and deserves further study.

Effect of repeated stress on a number of plasma amino acids and related compounds in young and old rats.

A number of amino compounds were studied in the plasma of young and old rats exposed to an immobilization stress on 3 different days. Baseline values were similar for all compounds between age groups except for slightly higher phenylalanine levels in old animals. Most differences were seen in the stress responses. Stress response intensities for alanine, aspartic acid, methionine, phenylalanine, and tyrosine were generally higher in old rats and recovery times from stress were usually longer. Although the stress responses decreased in young rats for some compounds over the three exposures, old rats showed no or less adaptation during this time. All the other amino acids showed no marked age effects. This study shows that certain plasma amino acids can serve as selective indicators of biological age and that their stress responses might be better markers of the aging process than their resting levels.

Effect of arginine administration on plasma and brain levels of arginine and various related amino compounds in the rat.

Arginine (ARG) was injected (0.8 g/kg, i.p.) into rats and levels of ARG were determined in plasma and four brain areas in the morning and afternoon. In control rats, brain values for ARG and some amino compounds are lower in the afternoon than in the morning. After ARG administration, ARG levels increase about 10-fold in the plasma and 2- to 3-fold in the brain areas. Brain ARG levels follow plasma levels. Elevated ARG levels affect a number of related amino compounds both in the plasma and all brain areas most notably ornithine, phosphoserine, glycine, GABA and ammonia. An increase of citrulline after ARG administration suggests the possibility of ARG-stimulated nitric oxide formation in the midbrain. Thus, ARG shows a daily rhythm in the plasma and brain and its administration increases ARG brain levels which seem to follow plasma levels. In addition, ARG alters a number of other amino compounds most notably GABA, glycine, ornithine and ammonia, indicating that some pharmacological effects seen after ARG administration might be caused by elevated levels of ARG and/or changes in other amino compounds.

Levels of taurine, amino acids and related compounds in plasma, vena cava, aorta and heart of rats after taurine administration.

A pharmacokinetic study on the fate of administered taurine in blood and some tissues and the effects on other amino compounds is presented. Injection of taurine (0.8 g/kg i.p.) causes markedly elevated plasma levels (70-fold at 15 min) which decrease later and approach baseline values after about 4 h. Concomitantly, other plasma amino compounds such as ornithine, threonine, asparagine, glutamine, alanine, citrulline, tyrosine, tryptophan, glycine, ammonia and arginine are reduced, whereas beta-alanine and phosphoserine are increased. At 30 min, tissue levels of taurine are roughly doubled in the vena cava and heart and tripled in the aorta. Other amino compounds affected are aspartic acid, serine, valine, methionine, tyrosine, ammonia, lysine, histidine, and arginine in the vena cava; aspartic acid, reduced glutathione, serine, and ammonia in the aorta; and reduced glutathione, alanine, citrulline and methionine in the heart. In most of these cases, plasma changes do not predict tissue changes which are generally substance- and tissue-specific. Thus, pharmacological effects seen after taurine administration could be caused by elevated taurine levels per se and/or taurine-induced changes in some of the amino acids and related compounds.

Effect of ethanol on amino acids and related compounds in rat plasma, heart, aorta, bronchus, and pancreas.

The composition of 36 amino acids and related amino compounds is presented for plasma, aorta, heart, pancreas, and bronchi. The distribution pattern of these biochemicals is similar but not identical among the tissues. The effect of an acute dose of ethanol (2 g/kg, i.p.) on these amino acids and related compounds was then studied. In the plasma, alanine, arginine, aspartic acid, beta-alanine, glycine, phenylalanine, and serine are decreased. In the aorta, ammonia and taurine are decreased. In the heart, aspartic acid and leucine are decreased, and carnosine, GABA, glutamic acid, and ornithine are increased. In the pancreas, asparagine and taurine are decreased, and citrulline, cysteine, histidine, and isoleucine are increased. In the bronchi, GABA, ethanolamine, histidine, taurine, and isoleucine are decreased. A number of correlations of some compounds in plasma or tissues were found but differed often between control and ethanol-treated animals. Ethanol-induced tissue changes generally do not correlate with plasma changes and are mostly specific for a given tissue.

The effects of arginine administration on the levels of arginine, other amino acids and related amino compounds in the plasma, heart, aorta, vena cava, bronchi and pancreas of the rat.

Arginine (0.8g/kg, ip) or a vehicle was administered to rats and the levels of arginine and a large number of related amino compounds++ were measured in plasma, heart, aorta, vena cava, pancreas and bronchi at specified time intervals. Arginine levels (nmol/ml) increased in the plasma from 237 to 3172 at 15 min, 1236 at 30 min and 509 at 120 min. Peak concentrations (nmol/g) of arginine are reached in the tissues at 15 or 30 minutes with control and postinjection values of 500 and 1769 in the heart, 314 and 1563 in the aorta, 575 and 2976 in the vena cava, 760 and 1943 in the bronchi, and 234 and 3638 in the pancreas. Arginine injection also affects a number of amino acids and related compounds in the plasma and tissues most notably ornithine, isoleucine, phosphoserine, leucine and ethanolamine. However, plasma level changes do not predict tissue level changes which are highly specific for an individual compound and tissue. There was no general indication that arginine injection stimulates nitric oxide (NO) formation in any tissue. Thus, arginine is rapidly absorbed from the abdominal cavity into the blood stream, is quickly taken up by the tissues studied and disappears after about 2 to 3 hours. The effects seen after arginine administration could be caused by arginine per se and/or changes in one or more of the related amino compounds but not by NO.

The effect of acute and chronic diazepam treatment on stress-induced changes in cortical dopamine in the rat.

The mesocortical dopamine system is thought to play an important role in the etiology of the stress response. Dopamine (DA) has been shown to accumulate in the rat frontal cortex in response to a wide variety of stressors. Diazepam, an anxiolytic benzodiazepine, can reverse the effects of stress on cortical DA. We investigated the effects of acute and chronic diazepam administration on immobilization stress-induced changes of the DA system in the frontal cortex of the rat. In the first study, 2.5 mg/kg diazepam was administered 20 min prior to 40 min of immobilization stress. Acute diazepam significantly reduced basal levels of extracellular DA and antagonized the stress-induced increase in cortical DA when compared to untreated stressed rats. Acute diazepam did not significantly effect extracellular DOPAC. In the second study, an experimental group of rats was given approximately 2 mg/kg/day diazepam in their drinking water for 3 weeks. This treatment significantly reduced anxiety as assessed by a staircase test for anxiety. Chronic diazepam had no effect on basal levels of cortical DA. However, chronic diazepam treatment also attenuated stress-induced increases in extracellular DA when compared to untreated stressed control rats. Chronic diazepam did not affect stress-induced changes in DOPAC but it did antagonize the effects of stress on HVA. Thus, acute and chronic diazepam treatment can antagonize stress-induced activation of the mesocortical DA system. It is proposed that this effect is produced through an enhancement of GABAergic neurotransmission by diazepam. The role of the dopaminergic system during stress, anxiety, and schizophrenia is discussed.

Effects of selective serotonergic agonists on aggressive behavior in rats.

The effects of the relatively specific serotonergic agonists 8-OH-DPAT (5-HT1A), TFMPP (5-HT1B), and DOB (5-HT2) were studied on defensive aggressive behavior in rats using the water competition test, 8-OH-DPAT (up to 0.25 mg/kg) and TFMPP (up to 1 mg/kg) were found to be ineffective, whereas DOB (up to 0.4 mg/kg) significantly reduced aggressive behavior in this test as well as in the offensive aggression test of the resident-intruder model. These results, combined with those from other studies, suggest that stimulation of 5-HT1A, 5-HT1B, and 5-HT2 receptors reduces offensive aggression, whereas defensive aggression is only decreased by 5-HT2 stimulation.