Membrane culture and reduced oxygen tension enhances cartilage matrix formation from equine cord blood mesenchymal stromal cells in vitro.

OBJECTIVE: Ongoing research is aimed at increasing cartilage tissue yield and quality from multipotent mesenchymal stromal cells (MSC) for the purpose of treating cartilage damage in horses. Low oxygen culture has been shown to enhance chondrogenesis, and novel membrane culture has been proposed to increase tissue yield and homogeneity. The objective of this study was to evaluate and compare the effect of reduced oxygen and membrane culture during in vitro chondrogenesis of equine cord blood (CB) MSC. METHODS: CB-MSC (n = 5 foals) were expanded at 21% oxygen prior to 3-week differentiation in membrane or pellet culture at 5% and 21% oxygen. Assessment included histological examination (H&E, toluidine Blue, immunohistochemistry (IHC) for collagen type I and II), protein quantification by hydroxyproline assay and dimethylmethylene assay, and mRNA analysis for collagen IA1, collagen IIA1, collagen XA1, HIF1α and Sox9. RESULTS: Among treatment groups, 5% membrane culture produced neocartilage most closely resembling hyaline cartilage. Membrane culture resulted in increased wet mass, homogenous matrix morphology and an increase in total collagen content, while 5% oxygen culture resulted in higher GAG and type II collagen content. No significant differences were observed for mRNA analysis. CONCLUSION: Membrane culture at 5% oxygen produces a comparatively larger amount of higher quality neocartilage. Matrix homogeneity is attributed to a uniform diffusion gradient and reduced surface tension. Membrane culture holds promise for scale-up for therapeutic purposes, for cellular preconditioning prior to cytotherapeutic applications, and for modeling system for gas-dependent chondrogenic differentiation studies.

The outcome of laparoscopic colorectal resection in T4 cancer.

AIM: Laparoscopic surgery for locally advanced tumours with extramural involvement is still controversial. It is believed that laparoscopic excision of T4 cancers is technically difficult and may result in prolonged operative time, increased conversion rate, added postoperative morbidity, and suboptimal oncological clearance. METHOD: Our unit has been practising laparoscopic colorectal surgery since 1992, and all data are entered into a database prospectively. Since 1999 we have routinely used the laparoscopic approach for colorectal cancer resections. Data regarding patients with a histologically T4 cancer operated on between 1999 and 2008 were analysed. Outcomes included operating time, conversion rate, postoperative complications and oncological outcome. RESULTS: Over a 10-year period, 146 patients (male 75) with a T4 cancer underwent laparoscopic resection. The median operating time was 125 (range, 46-285) min and the median blood loss was 50 (0-1800) ml. The conversion rate was 16%. Six (4.1%) patients experienced anastomotic leakage. The median number of lymph nodes harvested was 13 (2-40). One hundred and two (70%) patients underwent curative resection. The recurrence rates were 41% and 53% for stage II and III patients, respectively. Four (3.9%) patients had local recurrence. At a median follow up of 18 (1-118) months, the overall survival was 25 months, with median overall survival for patients with stage II, III and IV disease being 63, 36 and 12 months, respectively. CONCLUSION: Laparoscopic colectomy in histologically T4 cancer is safe. Oncological outcomes remain satisfactory. Based on our data, provided expertise is available, patients with locally advanced tumours should not be excluded from a laparoscopic approach.

Clb/Cdc28 kinases promote nuclear export of the replication initiator proteins Mcm2-7.

BACKGROUND: In the budding yeast Saccharomyces cerevisiae, the cyclin-dependent kinases of the Clb/Cdc28 family restrict the initiation of DNA replication to once per cell cycle by preventing the re-assembly of pre-replicative complexes (pre-RCs) at replication origins that have already initiated replication. This assembly involves the Cdc6-dependent loading of six minichromosome maintenance (Mcm) proteins, Mcm2-7, onto origins. How Clb/Cdc28 kinases prevent pre-RC assembly is not understood. RESULTS: In living cells, the Mcm proteins were found to colocalize in a cell-cycle-regulated manner. Mcm2-4, 6 and 7 were concentrated in the nucleus in G1 phase, gradually exported to the cytoplasm during S phase, and excluded from the nucleus by G2 and M phase. Tagging any single Mcm protein with the SV40 nuclear localization signal made all Mcm proteins constitutively nuclear. In the absence of functional Cdc6, Clb/Cdc28 kinases were necessary and sufficient for efficient net nuclear export of a fusion protein between Mcm7 and the green fluorescent protein (Mcm7-GFP), whereas inactivation of these kinases at the end of mitosis coincided with the net nuclear import of Mcm7-GFP. In contrast, in the presence of functional Cdc6, which loads Mcm proteins onto chromatin, S-phase progression as well as Clb/Cdc28 kinases was required for Mcm-GFP export. CONCLUSIONS: We propose that Clb/Cdc28 kinases prevent pre-RC reassembly in part by promoting the net nuclear export of Mcm proteins. We further propose that Mcm proteins become refractory to this regulation when they load onto chromatin and must be dislodged by DNA replication before they can be exported. Such an arrangement could ensure that Mcm proteins complete their replication function before they are removed from the nucleus.

Brain neurotransmitter turnover rates during rat intravenous cocaine self-administration.

The turnover rates of dopamine, norepinephrine, serotonin, aspartate, glutamate and GABA were measured in 27 brain regions of rats self-administering cocaine and in yoked cocaine- and yoked vehicle-infused controls using radioactive pulse-labeling procedures to identify brain neuronal systems underlying self-administration. Changes in the activity of heretofore unrecognized dopamine, norepinephrine, serotonin, glutamate and GABA innervations of the forebrain specific to cocaine self-administration were found. This included innervations of the nucleus accumbens, ventral pallidum, lateral hypothalamus and the anterior and posterior cingulate, entorhinal-subicular and visual cortices. Turnover rates also were calculated using metabolite/neurotransmitter ratios which were inconsistent with the pulse-label technologies indicating that ratio procedures are not accurate measures of neurotransmitter utilization. Results with the pulse-label technique provide evidence of the involvement of neuronal systems in cocaine self-administration not previously known, some of which may have a broader role in brain reinforcement processes for natural reinforcers (i.e. food, water, etc.) since drugs of abuse are thought to produce reinforcing effects by modulating activity in these endogenous systems.

Ventral pallidal extracellular fluid levels of dopamine, serotonin, gamma amino butyric acid, and glutamate during cocaine self-administration in rats.

RATIONALE: Dopamine innervation of the nucleus accumbens is thought to have a major role in the biological processes underlying cocaine self-administration. Recent data suggest that dopamine innervation of the ventral pallidum (VP) may also play an important role. OBJECTIVES: This experiment was initiated to assess extracellular fluid levels of dopamine (DA), serotonin (5-HT), gamma-aminobutyric acid (GABA), and glutamate (Glu) in the VP of rats self-administering cocaine using in vivo microdialysis. METHODS: Rats were implanted with intravenous jugular catheters and a microdialysis probe guide cannula into the VP and trained to self-administer (SA) three different doses of cocaine during each daily session. Other rats (yoked rats) were surgically prepared in identical fashion and received vehicle infusions during microdialysis sessions when the SA rat to whom they were yoked produced cocaine infusions. When stable baselines of self-administration were obtained, microdialysates were collected during two consecutive daily self-administration sessions. Neurotransmitter levels were measured using HPLC with electrochemical (DA and 5-HT) or fluorescence detection (GABA and Glu). RESULTS: In SA rats, extracellular fluid levels of DA [DA]e and 5-HT [5-HT]e were elevated throughout the session and levels of Glu [Glu]e showed small increases at a few isolated time points during the session. The increases in [DA]e and 15-HT]e were dose-dependent. Extracellular fluid levels of GABA [GABA]e were unchanged, as were levels of all four neurotransmitters in the yoked rats. CONCLUSIONS: These data support a potential role for DA and 5-HT innervations of the VP in intravenous cocaine self-administration.

Differences in extracellular dopamine concentrations in the nucleus accumbens during response-dependent and response-independent cocaine administration in the rat.

Studies indicate that nucleus accumbens (NAcc) dopamine neurotransmission is involved in the reinforcing and direct effects of cocaine. The present study was initiated to explore further the relationship of NAcc extracellular dopamine concentrations ([DA]e) and cocaine self-administration using a yoked littermate design. In the first experiment, one rat from each litter was trained to self-administer cocaine i.v. (SA: 0.33 mg/inf) under a fixed ratio 2 schedule, while a second rat received simultaneous infusions of cocaine yoked to the infusions of the SA (YC). NAcc [DA]e and cocaine concentrations ([COC]) were assessed during the test sessions using in vivo microdialysis combined with microbore HPLC procedures. [DA]e and [COC] were significantly elevated in the SA and YC groups during the self-administration session; however, [DA]e were greater in the SA group compared to the YC group in the first hour of the session, even though [COC] were not significantly different. On the following day, the rats previously allowed to self-administer cocaine were administered response-independent cocaine infusions yoked to the infusion pattern from the previous day. [DA]e were significantly elevated above baseline levels during the session but were significantly less than concentrations obtained when cocaine was self-administered by these subjects. [COC] during the sessions were not significantly different between the two days. Baseline [DA]e were not significantly different between the SA and YC groups or between Day 1 and Day 2. Furthermore, there was no significant difference in the in vitro probe recovery between one and two days following probe implantation. These results suggest that the context in which cocaine was administered significantly altered the neurochemical response to equivalent brain concentrations of cocaine. NAcc [DA]e was significantly increased when the delivery of cocaine infusions was contingent on the behavior of the rat, indicative of a role in the neural processes underlying cocaine reinforcement.

Rat brain neurotransmitter turnover rates altered during withdrawal from chronic cocaine administration.

This experiment utilized neurotransmitter turnover rates to assess the effects of withdrawal from chronic cocaine on the brain. A triad-littermate design was used to evaluate the importance of response dependency on the effects of withdrawal from chronic cocaine administration upon brain biogenic monoamine and amino acid putative neurotransmitter turnover rates. Each member of a triad was exposed to one of three conditions. Cocaine infusions (0.33 mg/inf) were used to engender and maintain lever pressing by one rat under an FR 2 schedule, while the second and third rats received simultaneous infusions of either cocaine or saline, respectively. After a minimum of 15 days exposure to the three treatment conditions and 24 h after the start of the last drug session, the triads were pulse labeled with [14C]glucose, [3H]tyrosine and [3H]tryptophan and killed 60 or 90 min later by total immersion in liquid nitrogen, The frozen brains were removed and dissected at -20 degrees C into 22 areas. The content and specific radioactivities for dopamine (DA), noradrenaline (NA), serotonin (5-HT), aspartate (Asp), glutamate (Glu), glycine (Gly) and gamma-aminobutyric acid (GABA) were determined in each brain region using high pressure liquid chromatography with electrochemical (biogenic monoamines) or fluorescence (amino acids) detection followed by liquid scintillation spectrometry. Turnover rates (TOR) were calculated and compared across treatment conditions. The significant decreases in TOR resulting from chronic cocaine exposure included 5-HT in the frontal cortex, DA in the cingulate cortex, entorhinal-subicular and motor-somatosensory cortices and NA in the inferior colliculus. Significant increases in TOR were also observed which included 5-HT in the preoptic-diagonal band region, DA in the hippocampus and NA in the pyriform and temporal-auditory cortices, the dentate gyrus and brainstem. GABA TOR was also increased in the preoptic-diagonal band region, dentate gyrus and brainstem of both groups receiving cocaine as was Glu TOR in the pyriform cortex and cerebellum. In addition, changes were seen in the rats under the ratio schedule of cocaine self-administration that were not seen in rats receiving yoked-cocaine infusions that included increased TOR of 5-HT in the pyriform cortex, NA in the caudate-putamen and GABA in the pyriform and motor-somatosensory cortices. Decreased 5-HT TO was seen in the motor-somatosensory cortex and dentate gyrus in the rats that had self-administered cocaine compared to the yoked-cocaine infused group. Perhaps the most interesting changes were those seen in the yoked-cocaine group that were reversed in the rats whose responding was maintained by cocaine.(ABSTRACT TRUNCATED AT 400 WORDS)

Specific effects of punishment on biogenic monoamine turnover in discrete rat brain regions.

Specific effects of punishment on the turnover rates of norepinephrine, dopamine and serotonin (5-HT) in brain regions were investigated in rats exposed to punishment. Two yoked controls were also used in an attempt to separate the non-specific effects of response rate, reinforcement density and direct effects of punisher (foot shock). Punished and unpunished littermate rats had similar response rates, and the reinforcement density was almost identical for both groups. A third group (yoked-shock rats) received food and shock independent of responding whenever these were given to the punished rats. When compared to the unpunished rats, changes in the monoamine turnover rates resulting from the punishment were similar to the effects of yoked-shock with respect to the direction of action in most cases (13 out of 17 changes). These changes may be related to non-specific effects of the shock. Four changes by the punishment were determined to be specific effects of the punishment since the yoked-shock had no effect or changed the turnover to the opposite direction. Among these, increase in 5-HT turnover rate in the frontal cortex (greater than 7-fold) was the largest change. These results and reported effects of drugs which act on serotonergic systems on the punished behavior suggest that the increase in 5-HT neuronal activity in the frontal cortex is involved in the behavioral suppression induced by the punishment.

Transient alterations in neurotransmitter levels during a critical period of neural development in coho salmon (Oncorhyncus kisutch).

The time of parr-smolt transformation (PST) in salmon is a critical period of brain development during which the olfactory imprinting on the natal stream takes place. PST is associated with a surge of plasma thyroxine. We report here similar surges in the brain content of serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), dopamine (DA), norepinephrine (NE), and glutamine (Gln). Other putative neurotransmitters, such as glutamate and aspartate show no such change while gamma-amino-butyric acid (GABA) shows a minor elevation.

Biogenic monoamine turnover in discrete rat brain regions is correlated with conditioned emotional response and its conditioning history.

The content and turnover of dopamine, norepinephrine and 5-hydroxytryptamine (serotonin), and the content of their respective major metabolites were evaluated in 19 discrete brain areas of rats exposed to conditioned emotional response (CER), and in control groups which received either equivalent yoked shock (shock only) or compound stimulus presentation (tone only). On test day, CER animals suppressed responding and exhibited forms of emotional behavior after presentation of the conditioned stimulus (CS); while shock only and tone only control groups, and CER animals which received an acute dose of diazepam prior to testing, did not suppress. Few changes were observed in content of the biogenic amines or their metabolites, suggesting that the behavioral manipulations were acting within normal physiological limits. On the other hand, numerous changes were observed in the utilization of the 3 biogenic monoamines, which were correlated with the conditioning-anxiety (comparisons of CER vs shock only) and the shock history (comparison of shock only vs tone only). These observations are consistent with putative neural pathways in the frontal cortex, septum, nucleus accumbens, amygdala, striatum, hippocampus and brain stem (which utilize specific monoamines), and with discrete brain areas which have been implicated in classical conditioning and CER-related phenomena. These observations suggest roles for biogenic monoamines in mediating or responding to the classical conditioning and emotional components of the paradigm.

Modifications of dopamine D1 receptor complex in rats self-administering cocaine.

Cocaine is spontaneously and experimentally self-administered and, when given repeatedly, it induces a stable form of sensitization to a previously assessed minimum active dose. In the present study, triads of rats chronically implanted with a jugular catheter were treated as follows: one animal was trained to self-inject cocaine, while the other two passively received either cocaine or saline whenever the self-administering rat completed the response requirement. After 30 days of stable responding, the animals were sacrificed and dopamine D1 receptor density and adenylyl cyclase activity were measured in different brain areas. Animals receiving cocaine (both self-administering and yoked) showed a down-regulation of dopamine D1 receptor number and of dopamine stimulated adenylyl cyclase activity in the nucleus accumbens, as compared to saline rats. In the olfactory tubercle, dopamine stimulated adenylyl cyclase activity appeared selectively and significantly down-regulated in self-administering animals.

Limbic acetylcholine turnover rates correlated with rat morphine-seeking behaviors.

Acetylcholine (ACh) turnover rates were measured in fourteen brain regions of rats intravenously self-administering morphine and in yoked-morphine and yoked-vehicle infused littermates to identify cholinergic neuronal pathways potentially involved in opiate reinforcement processes. Rats receiving chronic passive administration of morphine had increased ACh turnover rates in the frontal cortex and diagonal band and decreased rates in the medial septum. The significant changes in animals self-administering the drug were prominent in limbic regions with increases in the frontal cortex and decreases in the pyriform cortex, nucleus accumbens, amygdala and ventral tegmental area. Some components of opiate reinforcement may be mediated by increases in the activity of cholinergic ventral pallidal and diagonal band fibers innervating the frontal cortex and by decreases in activity of cholinergic fibers innervating the ventral tegmental area. These data and turnover rates for dopamine, norepinephrine, serotonin, aspartate, glutamate and gamma-aminobutyric acid previously determined in similarly treated animals are consistent with two neuronal circuits that may be involved in opiate seeking behaviors and opiate reinforcement processes.

Limbic muscarinic cholinergic and benzodiazepine receptor changes with chronic intravenous morphine and self-administration.

Muscarinic cholinergic and benzodiazepine receptor affinities and densities were evaluated in membranes from seven brain regions of rats intravenously self-administering morphine and in littermates receiving yoked-morphine or yoked vehicle infusions to identify neuronal systems potentially involved in mediating opiate reinforcement processes. Passive morphine infusion resulted in increases in muscarinic cholinergic receptor densities in the pyriform cortex and in decreases in the cingulate cortex while benzodiazepine receptor densities were decreased in both the hippocampal formation and entorhinal-subicular cortex compared to littermates receiving passive infusions of vehicle. Morphine self-administration resulted in decreased muscarinic cholinergic receptor densities in the frontal and entorhinal-subicular cortices and increases in the amygdaloid complex compared to littermates receiving yoked passive drug. These data are in agreement with acetylcholine turnover rate measurements in these animals and support the proposed role of cholinergic innervations of the frontal and entorhinal-subicular cortices and amygdaloid complex in opiate reinforcement processes.

Use of a single compartment LCEC cell in the determinations of biogenic amine content and turnover.

Content and specific radioactivity of the biogenic monoamines and content of their precursors and metabolites were simultaneously determined in CNS tissue extracts with a high pressure liquid chromatography system (HPLC). The content of dopamine, norepinephrine, serotonin. 5-hydroxyindoleacetic acid, homovanillic acid, 3,4-dihydroxyphenylacetic acid and 4-hydroxy-3-methoxyphenylethyleneglycol and the turnover rate of dopamine, norepinephrine and serotonin were measured in discrete rat brain regions using a one compartment electrochemical detector cell coupled to a C18-reverse phase HPLC column. The small fluid volume dead space of the cell allows the direct and precise collection of individual peaks for determining specific radioactivities. This method is especially suitable for central nervous system tissue samples from 8-20 mg wet weight and the sensitivity of the system in its routine configuration is approximately 2 pmol. This method for determinations of turnover is appropriate for investigations of animals in sensitive behavioral paradigms.U

Specific effects of punishment on amino acids turnover in discrete rat brain regions.

Specific effects of punishment on the turnover rates of aspartate (Asp), glutamate (Glu) and gamma-aminobutyric acid (GABA) in 14 brain regions were investigated in rats exposed to punishment. Two yoked controls were also used in an attempt to separate the nonspecific effects of response rate, reinforcement density and direct effects of punisher (foot shock). Punished and unpunished littermate rats had similar response rates, and the reinforcement density was almost identical for both groups. A third group (yoked-shock rats) received food and shock independent of responding whenever these were given to the punished rats. When compared to the unpunished rats, the punishment increased the turnover rates of the three amino acids in all brain regions examined except GABA turnover in the caudate-putamen and preoptic-diagonal band. The majority of these changes by the punishment were similar to the effects of the yoked-shock (yoked-shock versus unpunished), although the magnitude of increase by the punishment was mostly larger than that by the yoked-shock. Six changes by the punishment (increase in the turnover rates of Asp in the thalamus, Glu in the hypothalamus and GABA in the cingulate cortex, entorhinal-subicular cortex, dentate gyrus and hypothalamus) appeared to be the specific effects of punishment since the yoked-shock did not affect these parameters. These results suggest that the punishment caused a hyperexcitation of the amino acidergic neurons in the limbic systems, particularly those in Papez's circuit.

Combination operant conditioning--liquid nitrogen immersion chamber for studying neurotransmitter systems and behavior.

Studies of neurochemical events associated with behavior require a method of tissue fixation that is rapid and does not itself produce neurochemical changes. An apparatus is described that permits immediate immersion of an unrestrained behaving animal into liquid nitrogen. This method of tissue fixation has the greatest versatility for studying multiple neurotransmitter systems. In addition to the measurement of neurotransmitter content and turnover, investigation of neurotransmitter receptors, enriched nerve ending fractions and enzyme activities are possible. The operant conditioning-liquid nitrogen immersion chamber described here can be used for studying these neurotransmitter systems as they relate to animal's responding on operant schedules of reinforcement.

Brain neurotransmitter turnover correlated with morphine-seeking behavior of rats.

Neurochemical substrates of intravenous opiate self-administration were investigated in rats using littermate controls for vehicle and passive morphine infusion. The rates of turnover of the putative neurotransmitters, dopamine, norepinephrine, serotonin, gamma-aminobutyric acid, aspartate and glutamate were concurrently measured in eleven brain regions of rats intravenously self-administering morphine and yoked-morphine or yoked-vehicle infused littermates. The passive infusion of morphine resulted in significant changes in the rates of turnover of the biogenic monoamine and amino acid neurotransmitters in six brain regions with the caudate nucleus-putamen-globus pallidus showing the most changes. The contingent infusion of morphine resulted in changes in utilization rates that were generally greater in both magnitude and number than the effects of the drug itself. Twenty-nine significant changes were observed in the self-administering group with most changes occurring in limbic structures. The neurotransmitter turnover rate changes resulting from contingent administration suggest that the drug administration environment is an important factor that should be considered in studies of interactions between drugs and neuronal systems.

Effects of 5,7-dihydroxytryptamine lesions of the nucleus accumbens on rat intravenous morphine self-administration.

The role of serotonergic innervations of the nucleus accumbens in the processes maintaining intravenous morphine self-administration were assessed. Pairs of male rat littermates were implanted with intravenous jugular catheters and bilateral injection guide cannulae into the central medial nucleus accumbens, made physically dependent on morphine and then allowed to intravenously self-administer with continuous access. When stable baselines of drug intake were obtained (2-3 weeks), one of each pair received bilateral microinjections of vehicle and the other 5,7-dihydroxytryptamine (5,7-DHT) into the nucleus accumbens. Response independent infusions of morphine were delivered for 24 hours at the previous rate of self-injection and the animals were again allowed to self-administer while drug intake was monitored for thirteen days. The littermate pairs were then sacrificed by immersion in liquid nitrogen, the brains removed at -20 degrees C and frozen sections of the cannulae tract taken for histological assessment. The nucleus accumbens, anterior caudate nucleus and pyriform cortex were removed at -20 degrees C and biogenic monoamine content determined. The 5,7-DHT lesions resulted in a significant increase in drug intake and significantly decreased the content of serotonin (5-HT) and 5-hydroxyindoleacetic acid in the nucleus accumbens (-49% and -30%, respectively) and 5-HT in the anterior caudate nucleus (-14%) and pyriform cortex (-17%). Dose-effect relationships were assessed in four additional animals before and after similar bilateral 5,7-DHT lesions.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of 6-OHDA lesions of the central medial nucleus accumbens on rat intravenous morphine self-administration.

The function of dopaminergic innervations of the central medial nucleus accumbens in the processes maintaining intravenous morphine self-administration was assessed by lesioning with 6-OHDA and comparing drug intake with sham-vehicle treated littermates. Localized bilateral lesions of this structure resulted in significant increases in morphine intake shifting the dose-effect relationship to the right with twice the dose necessary to maintain prelesion rates of self-administration. Content of dopamine and dihydroxyphenylacetic acid was decreased in the nucleus accumbens after the lesion, but unchanged in the adjacent pyriform cortex and anterior caudate nucleus-putamen, while serotonin was significantly decreased in the pyriform cortex. High affinity uptake measurements also suggested nucleus accumbens dopaminergic and pyriform cortex serotonergic innervations to be affected by the lesion. The shift to the right in the dose effect relationship after the lesion suggests these neuronal systems to be excitatory to the processes mediating self-administration.

Lack of an effect of 6-hydroxydopamine lesions of the nucleus accumbens on intravenous morphine self-administration.

The neurotoxin, 6-hydroxydopamine (6-OHDA), has been used to selectively destroy dopamine containing neurons in discrete brain regions. Lesions of the nucleus accumbens with this neurotoxin decrease or eliminate cocaine and amphetamine self-administration and either increase or do not affect opiate self-administration in rats with unrestricted access to food and water. This study reports the effects of 6-OHDA lesions of the nucleus accumbens on responding maintained by food, water or morphine (3.3 mg/infusion). Six male rats with continuous access to three response levers were trained on a concurrent chained, fixed-ratio 1, fixed-ratio 9 schedule of reinforcer presentation. After stable patterns of responding were maintained by the three reinforcers, dose-effect curves for morphine were determined by substituting other doses of morphine or vehicle for 24-hour periods. Bilateral sham vehicle or 6-OHDA lesions of the nucleus accumbens were then completed and the effects of the lesion on food, water and morphine intake determined. Dose-effect evaluations were repeated after the lesion. The 6-OHDA lesions did not significantly affect responding maintained by food, water or morphine. The absence of an effect is most likely not the result of an insensitive baseline since other neurotoxin lesions produce long-term and selective decrements in morphine self-administration without affecting food and water responding. Like so many other manipulations, the magnitude of the effect that a neurotoxin lesion can exert on behavior may depend on the specific procedures that are used to maintain responding.