An update on the CNS actions of TRH and its analogs.

This brief review will discuss the recent literature on several of the central actions of TRH and its analogs. The most prominent of these actions include: (1) the arousal or analeptic effect in drug narcotized animals or in concussion models; (2) the reversal of cognitive deficits produced by various drugs or procedures, and (3) the improvement of several neurological deficits produced in animal models of spinal and/or cerebellar injury. The mediation of these TRH effects by neurotransmitters is discussed. While little has been published on the human neuropsychopharmacology of TRH, and especially of its analogs, the future holds considerable therapeutic promise for these interesting drugs.

Species differences in stereoselective hydrolase activity in intestinal mucosa.

PURPOSE: The aim of this study is to investigate species differences in the stereoselective hydrolysis for propranolol ester prodrugs in mammalian intestinal mucosa and Caco-2 cells. METHODS: Hydrolase activities for propranolol prodrugs and p-nitrophenylacetate in man (age: 51-71 years), the beagle dog (age: 4 years) and Wistar rat (age: 8 weeks) intestinal mucosa, and also in Caco-2 cells (passage between 60-70) were estimated by determining the rate of production of proparanolol and p-nitrophenol, respectively. RESULTS: The hydrolase activities for both propranolol prodrugs and p-nitrophenylacetate were in the order of man > rat >> Caco-2 cells > dog for intestinal microsomes, and rat > Caco-2 cells = man > dog for intestinal cytosol. Dog microsomes showed stereoselective hydrolysis for propranolol prodrugs, but not those from human or rat. Interestingly, both subcellular fractions of Caco-2 cells showed remarkable R-enantioselectivity except acetyl propranolol. Enzyme kinetic experiments for each enantiomer of butyryl propranolol in microsomes revealed that dog possesses both low and high affinity hydrolases. Both Km and Vmax values in rat were largest among examined microsomes, while Vmax/Km was largest in man. Finally, it was shown that the carboxylesterases might contribute to the hydrolysis of propranolol prodrug in all species by inhibition experiments. CONCLUSIONS: The hydrolase activities for propranolol prodrugs and p-nitrophenylacetate in intestinal mucosa showed great species differences and those in human intestine were closer to those of rat intestine than dog intestine or Caco-2 cells.

Species differences for stereoselective hydrolysis of propranolol prodrugs in plasma and liver.

Species differences and substrate specificities for the stereoselective hydrolysis of fifteen O-acyl propranolol (PL) prodrugs were investigated in pH 7.4 Tris-HCl buffer and rat and dog plasma and liver subfractions. The (R)-isomers were preferentially converted to propranolol (PL) in both rat and dog plasma with the exception of isovaleryl-PL in rat plasma, although the hydrolytic activities of prodrugs in rat plasma were 5-119-fold greater than those in dog plasma. The prodrugs with promoieties (C(=O)CH(R)CH3) based on propionic acid showed marked preference for hydrolysis of the (R)-enantiomers in plasma from both species (R/S ratio 2.5-18.2). On the other hand, the hepatic hydrolytic activities of prodrugs were greater in dog than rat, especially in cytosolic fractions. The hydrolytic activity was predominantly located in microsomes of the liver in rat, while the cytosol also contributed to hepatic hydrolysis in dog. Hepatic microsomal hydrolysis in dog showed a preference for the (R)-isomers except acetyl- and propionyl-PL. Interestingly, in rat liver all types of prodrugs with substituents of small carbon number showed (S)-preference for hydrolysis. The hydrolyses of (R)- and (S)-isomers of straight chain acyl esters in rat liver microsomes were linearly and parabolically related with the carbon number of substituents, respectively, while these relationships were linear for both isomers in dogs.

Intraseptal microinjection of beta-funaltrexamine blocked a microwave-induced decrease of hippocampal cholinergic activity in the rat.

Acute (45 min) exposure to pulsed (2 microseconds pulse width, 500 pulses per second) 2450-MHz microwaves at a power density of 1 mW/cm2 (whole body specific absorption rate 0.6 W/kg) microwaves caused a decrease in cholinergic activity in the hippocampus of the rat as measured by the sodium-dependent high-affinity choline uptake. Microinjection of beta-funaltrexamine (1 microgram) into the septum before microwave exposure blocked this effect. These data indicate that mu-opioid receptors in the septum mediate a microwave-induced decrease in cholinergic activity in the hippocampus and support our hypothesis that microwaves at a whole body SAR of 0.6 W/kg can activate endogenous opioids in the brain.

Involvement of D1 and D2 dopamine systems in the behavioral effects of cocaine in rats.

Cocaine (5-40 mg/kg, intraperitoneally) enhanced locomotion and rearing accompanied with head circling and body shaking. Although at 40 mg/kg typical stereotypy licking occasionally appeared, 40% of the rats died. At doses that did not affect physiologic locomotion and rearing, the D1-receptor antagonist SCH23390 but not D2 antagonist raclopride inhibited locomotion and rearing stimulated by cocaine (20 mg/kg). All behavioral responses of cocaine were abolished with increasing doses of raclopride and SCH23390. Sulpiride, a D2 antagonist, exerted a biphasic effect on locomotor activity (i.e., a low dose of sulpiride increased and a high dose decreased cocaine-induced locomotor activity). Sulpiride enhanced head circling, body shaking, and increases of rearing induced by cocaine. D2-receptor agonists quinpirole and bromocriptine inhibited these responses, presumably by activating the typical stereotyped behaviors such as sniffing at low doses, and licking and gnawing at high doses. The lowest dose of bromocriptine inhibited all behaviors induced by cocaine without producing typical stereotyped behaviors in itself. SK+F38393, a D1-receptor agonist, in combination with cocaine did not induce typical stereotype, which results in a synergistic effect of D1 and D2-receptor activities. The increases of locomotion and rearing, head circling, and body shaking induced by cocaine may involve the indirect activation of postsynaptic D1 and D2 receptors, presumably via dopamine release, resulting from inhibition of the presynaptic D2 receptors. These results also provide evidence that the indirect stimulation of postsynaptic D2 receptors by cocaine (20 mg/kg) is insufficient to induce stereotyped behaviors, and that the role of dopamine D1 receptors in mediating the behavioral actions of acute cocaine appears to be more important than that of D2 receptors. Our results also suggest that bromocriptine may be useful for the treatment of acute cocaine poisoning.

Effects of low-dose TRH on cognitive deficits in the ECT postictal state.

A low dose (0.5 mg) of thyrotropin-releasing hormone (TRH), a short-acting tripeptide with known analeptic properties, was administered to eight depressed patients 5 minutes after ECT session 3 or 4 in a double-blind, placebo-controlled crossover design. After TRH infusion the patients displayed selectively better performance on a battery of neuropsychological tests than they did after placebo infusion. Further exploration with pharmacological probes to mitigate ECT postictal cognitive deficits is warranted.

Microwave irradiation affects radial-arm maze performance in the rat.

After 45 min of exposure to pulsed 2450 MHz microwaves (2 microseconds pulses, 500 pps, 1 mW/cm2, average whole body SAR 0.6 W/kg), rats showed retarded learning while performing in the radial-arm maze to obtain food rewards, indicating a deficit in spatial "working memory" function. This behavioral deficit was reversed by pretreatment before exposure with the cholinergic agonist physostigmine or the opiate antagonist naltrexone, whereas pretreatment with the peripheral opiate antagonist naloxone methiodide showed no reversal of effect. These data indicate that both cholinergic and endogenous opioid neurotransmitter systems in the brain are involved in the microwave-induced spatial memory deficit.

The analeptic effect of methamphetamine in pentobarbital-narcotized rats is mediated via a dopaminergic-cholinergic mechanism.

Methamphetamine (MAP) administered in doses of 0.5 to 5 mg/kg i.p. to rats anesthetized with pentobarbital produced a shortening of the duration of loss of righting reflex. This analeptic effect of MAP was blocked by atropine but not by atropine methylbromide, indicating the central cholinergic nature of the response. This effect was also blocked by the D1 and D2 dopamine antagonists SCH 23390 (0.2 mg/kg) and raclopride (2 mg/kg), respectively. Pentobarbital decreased sodium-dependent high-affinity choline uptake (HACU) in frontal cortex and hippocampus as measured in synaptosomes from treated rats. MAP given to pentobarbital-narcotized rats restored HACU activity to nonanesthetized levels, but this restorative effect of MAP was blocked by SCH 23390 or raclopride. These data suggest that in addition to a cholinergic mechanism, the analeptic effect of MAP involves the dopamine system. alpha-Methyl-p-tyrosine, but not reserpine, pretreatment completely blocked the MAP analeptic response. In reserpinzed rats, MAP produced a markedly enhanced analeptic response. Studies of the effects of repeated administration of MAP on its analeptic activity were also undertaken in view of the well known sensitization to the locomotor and stereotypic effects of the amphetamines that occur with repeated intermittent administration. Rats pretreated daily with MAP (5 mg/kg) for 5 or 12 days showed neither tolerance nor sensitization to the analeptic effect of subsequent MAP administrations. 3H-quinuclidinyl benzilate-binding studies also showed no changes in muscarinic binding characteristics of membranes prepared from cortex or hippocampus of rats pretreated chronically with MAP. These and our earlier studies suggest that the analeptic effect of MAP is mediated via a dopaminergic-cholinergic mechanism.

The effects of SCH 23390 and raclopride on cocaine-induced analepsis and EEG arousal in rabbits.

The effects of the dopamine D1 and D2 receptor antagonists on cocaine-induced, cholinergically-mediated analeptic and hippocampal theta activity in anesthetized rabbits were investigated. Cocaine (2 mg/kg, i.v.) reduced by 35% the duration of loss of righting reflex produced by a 25 mg/kg dose of pentobarbital. This shortening of narcosis time was blocked by pretreating the animals with the D1 antagonist, SCH 23390 (0.1 mg/kg) but not with the D2 antagonist raclopride (1-2 mg/kg). Cocaine (5 mg/kg, i.v.) also produced a short burst of increased hippocampal theta activity in urethane-anesthetized rabbits, which was also blocked by SCH 23390 but not by raclopride. An unexpected finding was that raclopride itself, at 2 mg/kg (i.v.), produced a marked activation of theta activity that lasted for 15-20 min. When cocaine was administered after this time it produced a potentiated theta response, both in duration and in magnitude. These results suggest that in the rabbit, cocaine exerts a cholinergically-mediated behavioral and EEG arousal through a D1 dopamine mechanism. The role of the D2 system is less clear but appears to be antagonistic to the D1-mediated response.

Cocaine: evidence for NMDA- and opioid-mediated antinociception in the tail-flick test.

Cocaine (20-40 mg/kg, IP) produced in a rat tail-flick test a bimodal antinociceptive effect, the first peak appearing 5-30 min and the second 3-6 h after injection. The secondary, but not the initial, cocaine antinociception was blocked by naloxone (2 mg/kg, IP). The secondary antinociceptive action was also increased by a dose of the kappa-receptor agonist, trans-(+/-)-3,4-di-chloro-N-methyl-N-[2-(1-pyrrolidinyl) cyclohexyl]-benzene-acetamide methane sulfonate (U-50,488H) (5 mg/kg, IP), which by itself had no antinociceptive effect. Both the initial and secondary antinociceptions were increased following daily cocaine (20 mg/kg, IP) administration for 4 days. 3-(2-Carboxy-piperazin-4-yl) propyl-1-phosphonic acid (CPP) and (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine (MK-801), competitive and noncompetitive NMDA antagonists, respectively, inhibited both the initial and secondary cocaine antinociceptions. Although neither NMDA (5 mg/kg, IP) nor cocaine in lower doses (10 mg/kg, IP) alone produced antinociception, cocaine administered after NMDA produced antinociception. The dopamine D1 receptor agonist 1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazepin-7-ol (SK&F38393) enhanced both initial and secondary cocaine antinociceptions, while the D1 receptor antagonist R(+)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3-benzazepin -7-ol (SCH23390) reversed this potentiating effect. The D2 receptor agonist quinpirole and its antagonist sulpiride exerted opposite effects of their respective D1 counterparts. Both D2 agonist and antagonist were without activity by themselves.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of a 60 Hz magnetic field on central cholinergic systems of the rat.

We studied the effects of an acute (45 min) exposure to a 60 Hz magnetic field on sodium-dependent, high-affinity choline uptake in the brain of the rat. Decreases in uptake were observed in the frontal cortex and hippocampus after the animals were exposed to a magnetic field at flux densities > or = 0.75 mT. These effects of the magnetic field were blocked by pretreating the animals with the narcotic antagonist naltrexone, but not by the peripheral opioid antagonist, naloxone methiodide. These data indicate that the magnetic-field-induced decreases in high-affinity choline uptake in the rat brain were mediated by endogenous opioids in the central nervous systems.

Opioid receptor subtypes that mediate a microwave-induced decrease in central cholinergic activity in the rat.

We performed experiments to investigate subtypes of opioid receptors in the brain involved in the effect of acute (45 min) pulsed microwave exposure (2,450-MHz, 2-microseconds pulses, 500 pps, average power density 1 mW/cm2, peak-power density, 1 W/cm2, average whole body SAR 0.6 W/kg) on cholinergic activity in the rat brain. Rats were pretreated by microinjection of specific antagonists of mu, delta, and kappa opioid-receptors into the lateral cerebroventricle before exposure to microwaves. The data showed that all three subtypes of opioid receptors are involved in the microwave-induced decrease in cholinergic activity in the hippocampus. However, the microwave-induced decrease in cholinergic activity in the frontal cortex was not significantly affected by any of the drug treatments, confirming our previous conclusion that the effect of microwaves on the frontal cortex is not mediated by endogenous opioids.

Single vs. repeated microwave exposure: effects on benzodiazepine receptors in the brain of the rat.

We studied the effects of single (45 min) and repeated (ten daily 45-min sessions) microwave exposures (2450-MHz, 1 mW/cm2, average whole-body SAR of 0.6 W/kg, pulsed at 500 pps with pulse width of 2 microseconds) on the concentration and affinity of benzodiazepine receptors in the cerebral cortex, hippocampus, and cerebellum of the rat. We used a receptor-binding assay with 3H-flunitrazepam as ligand. Immediately after a single exposure, an increase in the concentration of receptor was observed in the cerebral cortex, but no significant effect was observed in the hippocampus or cerebellum. No significant change in binding affinity of the receptors was observed in any of the brain-regions studied. In rats subjected to repeated exposures, no significant change in receptor concentration was found in the cerebral cortex immediately after the last exposure, which may indicate an adaptation to repeated exposures. Our data also show that handling and exposure procedures in our experiments did not significantly affect benzodiazepine receptors in the brain. Because benzodiazepine receptors in the brain are responsive to anxiety and stress, our data support the hypothesis that low-intensity microwave irradiation can be a source of stress.

A possible 5-HT3 component of thyrotropin-releasing hormone-induced increases in gastric motility in developing rats.

Intracisternal injection of thyrotropin-releasing hormone (TRH) increases gastric motility primarily via a vagal cholinergic mechanism. However, a serotonergic (5-HT) component may also exist. Rats (7, 10, 14, and > or = 50 days of age) were anesthetized and gastric motility monitored via an extraluminal strain gauge. Following baseline, ICS 205-930 which blocks 5-HT3 and 5-HT4 receptors (0.01, 0.10, or 1.0 mg/kg) was administered intraperitoneally, then 30 min later intracisternal TRH (5 or 10 micrograms). ICS 205-930 0.1 and 1.0 mg/kg blocked TRH-induced motility in 7-day-old rats. Results support a 5-HT3 or 5-HT4 receptor contribution to TRH-induced gastric motility stimulation, and suggest that receptor expression is dynamic during development.

D-1 agonist, SKF 38393, but not a D-2 agonist, produces a cholinergically mediated analeptic effect in rabbits.

SKF 38393 (2-15 mg/kg, IV), but not quinpirole, shortened the duration of loss of righting reflex produced in pentobarbital-narcotized rabbits. This effect was blocked by atropine (2-5 mg/kg, IV), but not by atropine methylbromide, suggesting that a central cholinergic mechanism was involved. The analeptic effect was also blocked by SCH 23390 (0.1 mg/kg, IV) or raclopride (5 mg/kg, IV). These results indicate that SKF 38393 activates central cholinergic neurons, which in turn initiate the analeptic effect. However, the fact that raclopride also blocked the SKF 38393 analeptic effect, but quinpirole did not exert any analeptic effect, suggests that a D-1/D-2 modulation of cholinergic systems may be involved in the SKF 38393-induced analeptic effect. These results also support our earlier findings and view that cocaine-induced analeptic activity is mediated by a dopaminergic-cholinergic mechanism.

Naltrexone pretreatment blocks microwave-induced changes in central cholinergic receptors.

Repeated exposure of rats to pulsed, circularly polarized microwaves (2,450-MHz, 2-microseconds pulses at 500 pps, power density 1 mW/cm2, at an averaged, whole-body SAR of 0.6 W/kg) induced biphasic changes in the concentration of muscarinic cholinergic receptors in the central nervous system. An increase in receptor concentration occurred in the hippocampus of rats subjected to ten 45-min sessions of microwave exposure, whereas a decrease in concentration was observed in the frontal cortex and hippocampus of rats exposed to ten 20-min sessions. These findings, which confirm earlier work in the authors' laboratory, were extended to include pretreatment of rats with the narcotic antagonist naltrexone (1 mg/kg, IP) before each session of exposure. The drug treatment blocked the microwave-induced changes in cholinergic receptors in the brain. These data further support the authors' hypothesis that endogenous opioids play a role in the effects of microwaves on central cholinergic systems.

D1 agonist SKF 38393 antagonizes pentobarbital-induced narcosis and depression of hippocampal and cortical cholinergic activity in rats.

SKF 38393 (5 mg/kg), but not quinpirole, shortened the duration of loss of righting reflex produced in pentobarbital-narcotized rats. This effect was blocked by atropine (2 mg/kg), but not by atropine methylbromide, suggesting involvement of central cholinergic mechanisms. The analeptic effect was also blocked by SCH 23390 (0.2 mg/kg) or raclopride (2 mg/kg). SKF 38393 also increased sodium dependent high affinity choline uptake (HACU) in cortical and hippocampal synaptosomes that had been depressed by pentobarbital. SCH 23390 or raclopride prevented the SKF 38393 reversal of the depressed HACU, indicating that both D1 and D2 mechanisms were involved mediating the analeptic effect. These results provide neurochemical evidence that cortical and hippocampal D1-mediated cholinergic activation results in a behavioral arousal (analeptic) response. They also suggest that DA mechanisms may be involved in regulation of cortical and hippocampal cholinergic neurons.

Corticotropin-releasing factor antagonist blocks microwave-induced decreases in high-affinity choline uptake in the rat brain.

Acute (45-min) irradiation with pulsed low-level microwaves (2450-MHz, 2 microseconds pulses at 500 pps, average power density of 1 mW/cm2, whole-body average specific absorption rate of 0.6 W/kg) decreased sodium-dependent high-affinity choline uptake (HACU) activity in the frontal cortex and hippocampus of the rat. These effects were blocked by pretreating the animals before exposure with intracerebroventricular injection of the specific corticotropin-releasing factor (CRF) receptor antagonist, alpha-helical-CRF9-41 (25 micrograms). Similar injection of the antagonist had no significant effect on HACU in the brain of the sham-exposed rats. These data suggest that low-level microwave irradiation activates CRF in the brain, which in turn causes the changes in central HACU.