Krystyna M. Wozniak

Selective inhibition of NAALADase, which converts NAAG to glutamate, reduces ischemic brain injury

We describe here a new strategy for the treatment of stroke, through the inhibition of NAALADase (N-acetylated-α-linked-acidic dipeptidase), an enzyme responsible for the hydrolysis of the neuropeptide NAAG (N-acetyl-aspartyl-glutamate) to N-acetyl-aspartate and glutamate. We demonstrate that the newly described NAALADase inhibitor 2-PMPA (2-(phosphonomethyl)pentanedioic acid) robustly protects against ischemic injury in a neuronal culture model of stroke and in rats after transient middle cerebral artery occlusion. Consistent with inhibition of NAALADase, we show that 2-PMPA increases NAAG and attenuates the ischemia-induced rise in glutamate. Both effects could contribute to neuroprotection. These data indicate that NAALADase inhibition may have use in neurological disorders in which excessive excitatory amino acid transmission is pathogenic.

Focal application of alcohols elevates extracellular dopamine in rat brain: a microdialysis study

Dopaminergic systems are thought to play a major role in the stimulant and reinforcing properties of drugs of abuse, including ethanol. The present study describes the effects of local perfusion with ethanol (and other alcohols) on extracellular dopamine in the striatum and nucleus accumbens. Following the establishment of basal dopamine levels (2-3 h), various concentrations of ethanol in artificial CSF (0.01-10% v/v) were slowly perfused through a microdialysis probe. Each dose of ethanol was found to increase dopamine concentrations in both the striatum and nucleus accumbens. This increase was dose-related in the striatum. The exclusion of calcium and inclusion of 12.5 mM magnesium in the perfusion medium prevented, or greatly attenuated the ethanol-induced dopamine (DA) release. Thus, the release of DA by ethanol is exocytotic in nature and involves calcium-dependent processes. The other alcohols tested, namely methanol and butanol, demonstrated a structure-activity relationship together with ethanol, in their ability to increase extracellular DA. The relative potency was butanol greater than ethanol greater than methanol. The diffusion of ethanol into the brain tissue was investigated following perfusion through the probe. Relatively low concentrations of ethanol were found in striatal tissue during perfusion and they declined rapidly with time, following the removal of ethanol from the perfusate. The concentrations of ethanol achieved in brain tissue following focal application through the microdialysis probe were relevant to human intoxication.

Antagonism of 5-HT3 receptors attenuates the effects of ethanol on extracellular dopamine.

The effect of a 5-HT3 antagonist, ICS 205-930, on ethanol-induced changes in extracellular dopamine, was investigated with in vivo microdialysis. Pretreatment of rats with ICS 205-930 effectively attenuated the subsequent increases in dopamine, in both the nucleus accumbens and corpus striatum. This suggests that 5-HT may be involved in the effects of ethanol on dopaminergic systems.

Glutamate carboxypeptidase II inhibition protects motor neurons from death in familial amyotrophic lateral sclerosis models

Approximately 10% of cases of amyotrophic lateral sclerosis (ALS), a progressive and fatal degeneration that targets motor neurons (MNs), are inherited, and ≈20% of these cases of familial ALS (FALS) are caused by mutations of copper/zinc superoxide dismutase type 1. Glutamate excitotoxicity has been implicated as a mechanism of MN death in both ALS and FALS. In this study, we tested whether a neuroprotective strategy involving potent and selective inhibitors of glutamate carboxypeptidase II (GCPII), which converts the abundant neuropeptide N-acetylaspartylglutamate to glutamate, could protect MNs in an in vitro and animal model of FALS. Data suggest that the GCPII inhibitors prevented MN cell death in both of these systems because of the resultant decrease in glutamate levels. GCPII inhibition may represent a new therapeutic target for the treatment of ALS.

Food intake, neuroendocrine and temperature effects of 8-OHDPAT in the rat

Administration of 8-hydroxy-2(di-n-propylamino)tetralin (8-OHDPAT) to rats produced dose-dependent decreases in food intake and hypothermia, increases in plasma prolactin and corticosterone, and a decrease in plasma growth hormone. 8-OHDPAT administration also induced the serotonin behavioral syndrome at all doses. Pretreatment with metergoline did not affect the 8-OHDPAT-induced behavioral syndrome or decrease in food intake but attenuated the prolactin increase and, furthermore, potentiated 8-OHDPAT-induced hypothermia. Pretreatment with ritanserin or naloxone did not modify 8-OHDPAT-induced changes in food intake, temperature or prolactin. Similarly, pretreatment with phenoxybenzamine, propranolol, clonidine, haloperidol and methiothepin also did not attenuate 8-OHDPAT-induced decreases in food intake. Administration of pindolol alone produced hyperthermia, decreased food intake and enhanced prolactin secretion. Pindolol thus appears to act as a partial 5-HT agonist in addition to being an antagonist at central 5-HT receptors.

Behavior of streptozotocin-diabetic mice in tests of exploration, locomotion, anxiety, depression and aggression

The present study examined behavior of streptozotocin-diabetic mice in Porsolts swim test, a putative animal model of depression, in the holeboard test of exploration and locomotor activity, in the plus maze test of anxiety, and in the resident-intruder paradigm of aggression. Two weeks after an IP injection of 200 mg/kg streptozotocin, which caused a 20% weight loss and increased fluid consumption and urination, male NIH Swiss mice were found to show lengthened duration of immobility in the swim test. One week of insulin treatment (0.1 IU/g/day) partially antagonized this change. The locomotor activity scores in the streptozotocin-treated mice were lower in the holeboard but higher in the plus maze than in the controls; therefore, the lengthened immobility was not likely to be due to a general motor impairment. No significant changes in the time spent in social interaction or aggressive behavior were found in the streptozotocin-treated mice. The results indicate that streptozotocin-treated mice show lengthened immobility in the swim test.

The effect of 8-OH-DPAT on temperature in the rat and its modification by chronic antidepressant treatments

Administration of 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) to rats produced a dose-dependent hypothermia. Pretreatment with the receptor antagonist methiothepin abolished this effect, and pretreatment with haloperidol, propranolol and pindolol partially attenuated it, although methiothepin and pindolol had hyperthermic actions of their own. Other receptor antagonists including ritanserin, naloxone, clonidine, phenoxybenzamine and metergoline did not significantly modify the response elicited by subsequent 8-OH-DPAT challenge. In antidepressant studies, chronic treatment (22 days) with clorgyline attenuated the hypothermic response to 8-OH-DPAT, whereas similar duration of treatment with the tricyclics clomipramine and imipramine did not significantly modify it. Also, acute treatment for three days with each of the antidepressants did not modify 8-OH-DPAT-induced hypothermia. We conclude that rat rectal temperature can be a useful model to help assess the functional state of serotonergic mechanisms, including the adaptational changes induced by long-term antidepressant treatment.

Comparison of Neuropathy-Inducing Effects of Eribulin Mesylate, Paclitaxel, and Ixabepilone in Mice

Chemotherapy-induced neurotoxicity is a significant problem associated with successful treatment of many cancers. Tubulin is a well-established target of antineoplastic therapy; however, tubulin-targeting agents, such as paclitaxel and the newer epothilones, induce significant neurotoxicity. Eribulin mesylate, a novel microtubule-targeting analogue of the marine natural product halichondrin B, has recently shown antineoplastic activity, with relatively low incidence and severity of neuropathy, in metastatic breast cancer patients. The mechanism of chemotherapy-induced neuropathy is not well understood. One of the main underlying reasons is incomplete characterization of pathology of peripheral nerves from treated subjects, either from patients or preclinically from animals. The current study was conducted to directly compare, in mice, the neuropathy-inducing propensity of three drugs: paclitaxel, ixabepilone, and eribulin mesylate. Because these drugs have different potencies and pharmacokinetics, we compared them on the basis of a maximum tolerated dose (MTD). Effects of each drug on caudal and digital nerve conduction velocity, nerve amplitude, and sciatic nerve and dorsal root ganglion morphology at 0.25 × MTD, 0.5 × MTD, 0.75 × MTD, and MTD were compared. Paclitaxel and ixabepilone, at their respective MTDs, produced significant deficits in caudal nerve conduction velocity, caudal amplitude and digital nerve amplitudes, as well as moderate to severe degenerative pathologic changes in dorsal root ganglia and sciatic nerve. In contrast, eribulin mesylate produced no significant deleterious effects on any nerve conduction parameter measured and caused milder, less frequent effects on morphology. Overall, our findings indicate that eribulin mesylate induces less neuropathy in mice than paclitaxel or ixabepilone at equivalent MTD-based doses.

Evidence that 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI)-induced hyperthermia in rats is mediated by stimulation of 5-HT2A receptors

The effects of various 5-HT receptor subtype-selective antagonists were studied on phenylisopropylamine hallucinogen1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI)-induced hyperthermia in Wistar rats, in an attempt to characterize the 5-HT receptor subtype mediating DOI-induced hyperthermia. Intraperitoneal administration of DOI to rats produced hyperthermia with a peak effect at 60 min. Pretreatment with propranolol (β-adrenoceptor antagonist that also has binding affinity for 5-HT1A, 5-HT1B and 5-HT2C sites), MDL-72222 or ondansetron (5-HT3 antagonists) did not attenuate DOI-induced hyperthermia. In contrast, pretreatment with metergoline (5-HT1/5-HT2 antagonist), ketanserin, LY53857, mesulergine, mianserin and ritanserin (5-HT2C/5-HT2A antagonists), as well as spiperone (5-HT1A/5-HT2A/D2 antagonist), significantly attenuated DOI-induced hyperthermia. Furthermore, daily administration of DOI (2.5 mg/kg per day) for 17 days did not produce either tolerance to its hyperthermic effect or modifym-CPP-induced hyperthermia in rats. These findings suggest that DOI-induced hyperthermia in rats is mediated by stimulation of 5-HT2A receptors.

Differential neuroendocrine responses to the 5-HT agonist m-chlorophenylpiperazine in Fawn-Hooded rats relative to Wistar and Sprague-Dawley rats.

The effect of various doses of the 5-HT agonist m-chlorophenylpiperazine (MCPP) on neuroendocrine function (prolactin and corticosterone responses) were compared in three different rat strains: Wistar, Sprague-Dawley (SD), and Fawn-Hooded (FH) rats. Administration of various doses of MCPP produced increases in plasma concentrations of prolactin and corticosterone in all three rat strains. The prolactin responses of FH rats to MCPP were significantly smaller than that of either Wistar or SD rats, while corticosterone responses were equivalent across all three strains. On the other hand, baseline concentrations of corticosterone, but not of prolactin, were significantly higher in FH animals relative to both Wistar and SD animals. There was no significant difference in either baseline hypothalamic concentrations of serotonin, 5-hydroxyindoleacetic acid, norepinephrine, or dopamine or brain concentrations of MCPP among these three rat strains. These findings support some other data indicating that FH rats, a strain with a peripheral platelet serotonin storage pool disorder, also possess alterations in some neuroendocrine functions which are modulated by serotonin.