Interactions of thyrotropin releasing hormone and morphine sulfate in rodents.

Thyrotopin releasing hormone (TRH) produces “wet dog shakes” in rats similar to those observed during morphine withdrawal. The shaking behavior precipitated by morphine abstinence can be exacerbated by TRH administration while the other components of the morphine withdrawal syndrome remain unchanged. Morphine, chlorpromazine, apomorphine, and Δ⁹-tetrahydrocannabinol effectively block shakes induced by either TRH administration or morphine withdrawal. These results suggest the possibility that endogenous TRH may be associated with the “wet dog shakes” observed as a portion of morphine's abstinence syndrome in rats. However, TRH is unable to alter the stereospecific binding of morphine $\text{in}$$\text{vivo}$ or $\text{in}$$\text{vitro}$, and naloxone fails to potentiate the number of TRH-induced shakes. TRH has no antinociceptive properties, and it cannot alter those of morphine. These data suggest that more than one neuromechanism may be responsible for shaking behavior in rats.     

The effects of narcotic analgesics and narcotic antagonists on ganglionic transmission in the rat.

The effects of narcotic analgesics, narcotic-antagonist analgesics and narcotic antagonists on ganglionic transmission in the superior cervical ganglia of the rat were studied $\text{in}$$\text{vivo}$ and $\text{in}$$\text{vitro}$. $\text{In}$$\text{vivo}$ administration of morphine, meperidine, methadone, pentazocine or naltrexone blocked ganglionic transmission. Levorphanol, cyclazocine, nalorphine and naloxone had no effect on ganglionic transmission in this procedure. $\text{In}$$\text{vitro}$ studies confirmed the $\text{in}$$\text{vivo}$ results with the exception of levorphanol, cyclazocine and nalorphine, which were also found to block ganglionic transmission $\text{in}$$\text{vitro}$. In both preparations, naloxone did not antagonize the effect of morphine, suggesting that the effects of morphine and the other opiates were nonspecific. Similar potency of $\text{d}$- and $\text{l}$-isomers of pentazocine and cyclazocine support this conclusion. The observation that naltrexone blocked ganglionic transmission, but the other pure narcotic antagonist, naloxone, was inactive is somewhat unique to this test procedure and possibly significant.     

Effects of morphine on dopamine-stimulated adenylate cyclase and on cyclic GMP formation in primate brain amygdaloid nucleus.

In homogenates of $\text{Macaca}$$\text{mulatta}$ (Rhesus) or $\text{Cebus}$$\text{apella}$ amygdaloid nuclear complex, adenylate cyclase activity was approximately doubled by either 10μ$\text{M}$ dopamine or 8m$\text{M}$ NaF. In the presence of morphine, the stimulation by dopamine was reduced. A 90–100% inhibition of the dopamine stimulation was obtained with 20μ$\text{M}$, and a 50% inhibition, with 5μ$\text{M}$ morphine. The effects of 10μ$\text{M}$ morphine on dopamine stimulation were reversed by 10μ$\text{M}$ naloxone. Morphine itself did not significantly affect the basal adenylate cyclase activity, but in the presence of 10μ$\text{M}$ morphine the stimulation by 8m$\text{M}$ NaF was reduced approxiamtely 50%. The data suggest an action of morphine at a receptor site which is distinct from the dopamine receptor, but which inhibits the dopamine-stimulated adenylate cyclase. In addition, the cyclic GMP content of $\text{Cebus}$ amygdala slices was reduced by 50–75% during incubation for 5–20 minutes with morphine. Maximum effects on cyclic GMP were obtained with 10μ$\text{M}$, and half-maximum effects, with 0.1μ$\text{M}$ morphine. The effect of morphine on amygdala cyclic GMP was not reversed by naloxone. Thus, this action of morphine may not be receptor mediated, or may involve the interaction of morphine with receptors other than the opiate receptor.     

Mechanism of development of tolerance to injected morphine by guinea pig ileum.

Injection of a large dose of morphine into a guinea pig results in a block of electrically-induced contractions of the ileum $\text{in vitro}$. A similar dose is almost ineffective in guinea pigs given morphine chronically. The time course for development of this tolerance has been determined in guinea pigs injected twice daily with morphine 100 mg/kg and challenged on various days with 750 mg/kg of the drug. Animals similarly injected but not challenged served as controls. The inhibitory effect of the challenging dose on electrical stimulation of longitudinal muscle decreased with successive days of morphine administration; by the 10th day there was almost complete tolerance to the challenging dose. Sensitivity of the tissues of chronically morphinized unchallenged controls towards acetylcholine, serotonin, histamine and norepinephrine was essentially the same as that of naive animals. The potency of morphine $\text{in vitro}$ in blocking electrical stimulation was also unchanged by chronic morphine administration in the above manner. Thus tolerance to injected morphine cannot be explained by reduced affinity of the drug for the opiate receptor. Tissues of chronically morphinized animals gave a contracture with naloxone, the extent of the contracture increasing with time of drug administration. This naloxone effect is attributed to displacement of morphine from a new opiate receptor site induced during morphine administration. It is suggested that this new receptor is involved in tolerance to injected morphine as well as some aspects of the withdrawal syndrome.     

Actions of morphine on prostate gland fructose metabolism

Varying doses of morphine sulfate (10, 20 or 40 mg/kg daily × 10) were observed to suppress metabolic activities in the mouse prostate gland. Prostate gland fructose, an index of androgenic activity, was significantly reduced by these dose regimes of morphine (P < 0.01). Injections of morphine sulfate (20 mg/kg daily × 10) led to an inhibitition in the $\text{in vitro}$ synthesis of both fructose^?14C and sorbitol^?14C from glucose^?14C by the prostate gland, part of which may have been due to decreased uptake of glucose by the gland. The $\text{in vitro}$ assimilation of 2-deoxyglucose^?14C by the prostate was also reduced by morphine treatment. The $\text{in vitro}$ actions of morphine (2 × 10^?3M) on the metabolism of radioactive glucose by the mouse prostate gland likewise revealed a significant reduction in the formation of sorbitol^?14C, but no decrease in fructose^?14C formation. These results indicate that both the $\text{in vitro}$ and $\text{in vivo}$ actions of morphine can inhibit fructose metabolism in the prostate gland.     

Motility of the N-terminal tail of phosphorylase b as revealed by crosslinking.

There are lysyl-ε-NH₂ groups within about 3.5 Å distance across the intersubunit contact area of rabbit muscle phosphorylase $\text{b}$, as shown by cross-linking with malonic diimidate. These include the lysines of N-terminal region as revealed by limited tryptic digestion, but the contribution of the tail lysines to overall formation of covalent dimers is small. The fine structure of dimer band on dodecylsulfate-gelelectrophoretograms of crosslinked phosphorylases suggests that the tail retains its freedom in the phosphorylase $\text{b}$-AMP complex. Amidination induces the dissociation of phosphorylase $\text{b}$ dimer, which is slow relative to crosslinking.     

Inhibition of drug metabolism by chronically administered naltrexone

Naltrexone, a long-acting narcotic antagonist, was administered to mice via aong-term delivery system of 1.5 mm beads containing 2 mg of naltrexone in a 90/10 polylactic/glycolic acid copolymer (Dynatech R/D Comp.). A single bead implanted subcutaneously antagonized the analgesic action of interimittently administered morphine sulfate (20 mg/kg, i.p.) for 25–35 days. During this 4–5 week period during which the naltrexone was pharmacologically active, the activities of the hepatic, microsomal mixed function oxidases aminopyrine N-demethylase and aniline hydroxylase were depressed to 30–50% of the levels seen in sham-implanted controls. Hexobarbital sleeping time and zoxazolamine paralysis time were significantly prolonged, and the blood half-lives of ¹⁴C-pentobarbital and ¹⁴H-amphetamine were lengthened when the monoxygenase activities were inhibited. Sleeping time following administration of ethanol was unaffected. $\text{In}$$\text{vitro}$, both naltrexone and its major metabolite, ß-naltrexol, were found to be inhibitory of the activities of aminopyrine N-demethylase and aniline hydroxylase, although the parent compound was the more potent inhibitor of both activities by a factor of 2–3.     

Dexamethasone blocks morphine-induced hypothermia in restrained rats

Rats restrained in small plexiglass restraining cages responded to morphine, 30mg/kg, with a pronounced $\text{hypo}$thermia. However, when this same dose of morphine was administered to unrestrained freely moving rats a marked $\text{hyper}$thermia resulted. The absolute magnitude and time course for body temperature changes after morphine were similar in both groups. When restrained rats pretreated with dexamethasone were administered morphine the hypothermic response was not only prevented but a subsequent hyperthermia occured. Morphine injected into unrestrained dexamethasone pretreated rats, however, still elicited a hyperthermia although of slightly lesser magnitude. These findings indicate that restraint is a potent modifier of morphine's effects on thermoregulation and that this effect most probably results from a stress-related activation of anterior pituitary hormone release. The possibility that ACTH or ß-endorphin released by the stress of restraint are responsible for this modification is discussed.     

The inhibition of morphine: UDP-glucuronyltransferase in rabbit liver microsomes by cyproheptadine.

The reactivity of the non-narcotic substances, cyproheptadine and N-desmethylcyproheptadine with morphine: UDP-glucuronyltransferase was studied in rabbit hepatic microsomal preparations. Cyproheptadine produced a potent competitive inhibition of morphine glucuronidation $\text{in vitro}$ (K_i=0.08 mM) whereas its N-desmethyl derivative was significantly less effective (K_i=0.4 mM). No cyproheptadine glucuronide was formed in these reactions suggesting that cyproheptadine acts as a dead-end inhibitor. Results indicate that the mechanism of the inhibition of morphine: UDP-glucuronyltransferase by cyproheptadine is similar to that produced by opioids and is related to the presence of the N-alkyl group in it structure.     

Effect of morphine sulfate on adenylate cyclase and phosphodiesterase activities in rat corpus striatum.

The effect of morphine sulfate (MS) on adenylate cyclase (AC) and phosphodiesterase (PDE) activities in the rat striatum was investigated. MS produced a dose-dependent increase in basal AC activity and did not alter sodium fluoride-induced stimulation both $\text{in}$$\text{vivo}$ (7.5–30 mg/kg, 1 hr pretreatment, i.p.) and $\text{in}$$\text{vitro}$ (1–100μM). $\text{in}$$\text{vitro}$, when submaximal effective concentrations of dopamine and MS were combined, there was an additive effect. However, administration of MS $\text{in}$$\text{vivo}$ did not alter dopamine-induced stimulation of AC activity. MS, $\text{in}$$\text{vitro}$ and $\text{in}$$\text{vivo}$ inhibited PDE activity in a dose-dependent manner only with the high substrate concentration (3.3 × 10⁻³M cyclic AMP). Preliminary results from this study indicate that morphine affects the cyclic AMP system.     

Naloxone antagonism of phenoxybenzamine antinociception in the mouse tail stimulation test.

Phenoxybenzamine was antinociceptive in the mouse tail electrical stimulation assay (ED50, 36.8 mg/kg) with a peak effect at $\text{2}\text{1}\text{2}$ hours after subcutaneous injection. Naloxone antagonized this antinociception action of phenoxybenzamine in a dose-related manner. Dose-ratio analysis of naloxone's antagonism of phenoxybenzamine antinociception gave a pA₂ value of 6.15, similar to that found for the benzomorphinan mixed agonist-antagonists. This is in agreement with the sodium response ratio found for phenoxybenzamine, 4.3, in $\text{in vitro}$ assays of phenoxybenzamine inhibition of ³H-naloxone binding to mouse brain homogenate (5). These findings suggest that phenoxybenzamine binds to the opiate receptor both $\text{in vivo}$ as well as $\text{in vitro}$ in a manner similar to the mixed agonist-antagonists.     

Location and characterization of opiate receptors regulating pituitary secretion

The site at which opiate agonists and antagonists act to alter secretion of prolactin, growth hormone and luteinizing hormone as well as the pharmacological specificity of the opiate receptors mediating these effects were examined in rats. Injection of β-endorphin but not a 10 fold higher dose of the non opiate peptide β-endorphin, increased release of prolactin and growth hormone in male rats while inhibiting luteinizing hormone release in ovariectomized, estrogen primed female rats. Prior treatment with naltrexone i.p. blocked these responses. Injection of naltrexone into the hypothalamus lowered prolactin release. In rats with a surgically formed hypothalamic island systemic administration of morphine or naltrexone altered prolactin release in the same manner as was observed in intact animals. In contrast no effects of β-endorphin or naltrexone were observed on the spontaneous secretion of prolactin $\text{in}$$\text{vitro}$. In addition β-endorphin did not alter the inhibition of prolactin release produced by apomorphine $\text{in}$$\text{vitro}$. The ED₅₀ for stimulation of prolactin release following intraventricular administration of β-endorphin or the synthetic enkephalin analog FK 33-824 was the same, approximately 0.1 ng/rat. However FK 33-824 at 0.2 ng/rat was able to produce much greater analgesia and catatonia than β-endorphin. The metabolism and distribution of β-endorphin was examined but did not account for these differential effects. These results indicate that opiate agonists and antagonists can act at the hypothalamic but not the anterior pituitary level to alter release of prolactin, growth hormone and luteinizing hormone. In addition the data suggest that the opiate receptors mediating release of prolactin may have a different pharmacological specificity from those involved with analgesia and catatonia.     

Differential effect of environmental temperature on morphine physical dependence and abstinence

1) Ambient temperature (T_a) significantly influenced the display of 4 of the 14 naloxone-precipitated withdrawal signs (nesting, flat posture, vocalization, dyspnea) in morphine-dependent, non-hibernating ground squirrels ($\text{Citellus}$$\text{lateralis}$).2) Analysis of variance performed on the six quantified signs revealed that T_a during withdrawal, but not during the development of physical dependence, was a significant factor in determining the expression of two signs (nesting and vocalization).3) The interaction between the influence of T_a during the periods of morphine administration and abstinence was a significant factor in determining the expression of nesting behavior, a finding that is consistent with the natural role of nesting as a behavioral thermoregulatory response.4) We conclude that environmental temperature modulates the expression of selected components of the naloxone-precipitated abstinence syndrome in $\text{C. lateralis}$ without exerting a measurable influence on the development of morphine physical dependence itself.     

Ribonucleotide reductase activity in green algae

A ribonucleoside diphosphate reductase is demonstrated in the algae, $\text{Scenedesmus}\text{obliquus}$ and $\text{Chlorella}\text{pyrenoidosa}$. In synchronized cultures an activity maximum at the 12th hour of the cell cycle coincides with maximum DNA production. Induction of reductase activity is prevented by cycloheximide. The enzyme requires dithiols for reduction of CDP $\text{in}\text{vitro}$; it is not significantly stimulated by iron or magnesium ions nor dependent upon deoxyadenosylcobalamin. ATP stimulates the reaction but dATP or dTTP act as inhibitors. The ribonucleotide reductase of green algae differs from the B₁₂-requiring enzyme characterized in $\text{Euglena}\text{gracilis}$.     

Effects of morphine on one-trial appetitive learning

Rats given intraventricular injections of morphine before and after a single training trial on a water-finding task performed significantly better on a retention test 24 hours later than rats given Ringer's injections instead of morphine either before $\text{or}$ after the training trial, and significantly better than rats given Ringer's before and after the training. Tial. The effect of morphine was interpreted as a retroactive strengthening of associations that the rats had formed during the training trial. The phenomenon suggests a possible model for understanding certain positively reinforcing actions of morphine.     

Self-administration of heroin, acetylmethadol, morphine, and methadone in rhesus monkeys.

Heroin, α-$\text{l}$-acetylmethadol (LAAM), morphine, and methadone each maintained self-administration in rhesus monkeys. The order of relative potency was heroin ≥ LAAM > morphine ≥ methadone. Total daily drug intake increased as dose per injection increased; maximum daily intake was inversely related to relative potency. At high doses, self-injection of methadone and LAAM caused stupor and/or respiratory failure in some monkeys. These toxic effects were partly or completely reversible by naloxone.     

Evidence for morphine and morphine-like alkaloid responses resistant to naloxone blockade in the rat vas deferens.

The application of morphine or surrogates to the isolated rat vas deferens maintained at 37° C in Tyrode solution, produced an increase in the electrically induced muscular twitch. In contrast, leucine enkephalin or D-alanine₂methionine enkephalinamide produced a dose-dependent inhibition of the muscular twitch. The effect of morphine and derivatives was not antagonized by naloxone, but the depression caused by the opiate pentapeptides or β-Endorphin was readily antagonized and reversed by naloxone. Tolerance developed to the $\text{in vitro}$ effect of morphine; vasa deferentia obtained from tolerant-dependent rats were about six times less sensitive to the effect of morphine and about five times less sensitive to the depression caused by leucine enkephalin as compared to their respective paired, placebo implanted control rats.     

Smooth specific phage adsorption: endorhamnosidase activity of tail parts of P22

The tail parts of phage P22 as well as the phage particles cleave the O-antigen of its host bacterium, $\text{Salmonella typhimurium}$. The cleavage is caused by specific breakage of α-rhamnosyl 1–3 galactose linkages. Thus the tail parts of this phage consist of an enzyme, endorhamnosidase. The enzyme was not detected in nonpermissible strain infected with an amber gene 9 mutant of P22. Head without tail parts gains infectivity only after incubation with the tail parts which carry this enzymatic activity.     

Relationship between chronic treatment with morphine and sex-dependent low molecular weight protein excretion

Chronic administration of morphine caused a marked decrease in the urinary excretion of low molecular weight proteins (LMWP) including α₂u-globulin (E.L. Stanley and O.W. Neuhaus, Biochim. Biophys. Acta, $\text{257}$, 461–470, 1972), which exist only in male rats. Levels of urinary high molecular weight proteins, such as albumin and γ-globulin, however, were not significantly changed by morphine treatment. The amount of excreted LMWP was observed to decrease 3 days after giving morphine, 0.5 mg/g food, and to recover to the control level within 6 days after withdrawal of morphine. These findings suggest that the decrease of excreted LMWP may be related to the development of morphine dependence in male rats.