Opioids and behavior: genetic aspects

Three animal models, based on genetic differences in endogenous opioid peptides and opioid receptors, are described. Obese mice and rats, whose pituitary opioid content is elevated, may be used to investigate eating disorders. Recombinant inbred strains of mice, which differ in brain opioid receptors and analgesic responsiveness, can be used for study of opioid- and nonopioid-mediated mechanisms of pain inhibition. Individual reactivity to opioids can be examined in C57BL/6 and DBA/2 inbred strains of mice. A model that combines a variety of opioid effects is offered and suggests the existence of a genetically determined dissociation of opioid effects on locomotor activity and pain inhibition. In addition, stimulatory locomotor responses in the C57BL/6 reaction type are linked to a high risk of drug addiction and facilitatory effects on adaptive processes, while high analgesic potency in the DBA/2 reaction type is accompanied by a low proneness to drug abuse and amnesic properties of opioids.     

Genetic control of hippocampal cholinergic and dynorphinergic mechanisms regulating novelty-induced exploratory behavior in house mice

Summary Neurobehavioral genetics endeavors to trace the pathways from genetic and eenvironmental determinants to neuroanatomical and neurophysiological systems and, thence, to behavior. Exploiting genetic variation as a tool, the behavioral sequelae of manipulating these neuronal systems by drugs and antisera are analyzed. Apart from research in rats, this paper deals mainly with the genetically-influenced regulation in mice of exploratory behaviors that are adaptive in novel surroundings and are hippocampally-mediated. Special attention is paid to neuropeptidergic, GABAergic, and cholinergic synaptic functions in the mouse hippocampus.The behaviorally different inbred mouse strains C57BL/6 and DBA/2 show opposite reactions (reductions and increases, respectively, in exploration rates) to peripheral and intrahippocampal injections with agents that interfere with peptidergic, cholinergic, and GABAergic neurotransmission. These findings can be explained by an interdependent over-release of opioids, arrested GABA release, and excess acetylcholine in the hippocampal neuronal network of DBA/2 mice, as compared to C57BL/6 mice where these systems are functionally well balanced. Very similar results have been obtained with the lines SRH and SRL, derived from C57BL/6 and DBA/2, and genetically selected for rearing behavior. Most probably, the opioids act to disinhibit exploratory responses. An additional genetic approach is mentioned, in which four inbred mouse strains and one derived heterogeneous stock are used for estimating genetic correlations between structural properties of the hippocampal mossy fibers and levels of hippocampal dynorphin B, on the one hand, and frequencies of exploratory responses to environmental novelty, on the other.     

Genetic control of hippocampal cholinergic and dynorphinergic mechanisms regulating novelty-induced exploratory behavior in house mice

Neurobehavioral genetics endeavors to trace the pathways from genetic and environmental determinants to neuroanatomical and neurophysiological systems and, thence, to behavior. Exploiting genetic variation as a tool, the behavioral sequelae of manipulating these neuronal systems by drugs and antisera are analyzed. Apart from research in rats, this paper deals mainly with the genetically-influenced regulation in mice of exploratory behaviors that are adaptive in novel surroundings and are hippocampally-mediated. Special attention is paid to neuropeptidergic, GABAergic, and cholinergic synaptic functions in the mouse hippocampus. The behaviorally different inbred mouse strains C57BL/6 and DBA/2 show opposite reactions (reductions and increases, respectively, in exploration rates) to peripheral and intrahippocampal injections with agents that interfere with peptidergic, cholinergic, and GABAergic neurotransmission. These findings can be explained by an interdependent over-release of opioids, arrested GABA release, and excess acetylcholine in the hippocampal neuronal network of DBA/2 mice, as compared to C57BL/6 mice where these systems are functionally well balanced. Very similar results have been obtained with the lines SRH and SRL, derived from C57BL/6 and DBA/2, and genetically selected for rearing behavior. Most probably, the opioids act to disinhibit exploratory responses. An additional genetic approach is mentioned, in which four inbred mouse strains and one derived heterogeneous stock are used for estimating genetic correlations between structural properties of the hippocampal mossy fibers and levels of hippocampal dynorphin B, on the one hand, and frequencies of exploratory responses to environmental novelty, on the other.     

Different action of alpha-methyl-DOPA on sleep and labyrinth learning in 2 inbred strains of mice]

In two inbred mice strains C 57 BR and C 57 BL/6 presenting the same type of sleep, but a different capacity of learning, Alpha-Methyl-Dopa (100 mg/kg) injected after every session, suppresses paradoxical sleep completely for 9 to 11 h. Maze-learning performance is retarded in C 57 BR mice, but facilitated in C 57 BL/6.     

Arylsulfatase B synthesis and clearance in inbred mouse strains

Arylsulfatase B activity levels were approximately 2-3-fold higher in adult C57BL/6J liver and kidney compared to corresponding tissues from A/J inbred mice. In vivo incorporation of tritiated leucine into C57BL/6J hepatic arylsulfatase B reached a maximum approximately 15 h after injection. The label was cleared from C57BL/6J arylsulfatase B with an apparent half-life of 36 h. The relative rates of synthesis of C57BL/6J and A/J arylsulfatase B were similar; however, the A/J enzyme was cleared more rapidly from liver tissue. C57BL/6J kidney arylsulfatase B appeared to be synthesized at a 2-3-fold higher rate than the corresponding A/J enzyme. These trends suggest genetic regulation of arylsulfatase B is effected through different means in liver and kidney from adult mice of these two inbred strains.     

Genetic differences between inbred strains of mice; a new source of variation in high-sulphur keratins

Proteins were extracted by improved techniques from the hair of inbred strains of Peru, C57BL/6 and CBA/Ca mice. The extracts were characterized by amino acid analysis and high resolution polyacrylamide gel electrophoresis, and previously unreported strain differences were observed. Genetic analysis confirmed them.     

Natural peptide analgesics: the role of solution conformation

Endogenous opioids have been studied extensively since their discovery, in the hope of finding a perfect analgesic, devoid of the secondary effects of alkaloid opioids. However, the design of selective opioid agonists has proved very difficult. First, structural studies of peptides in general are hampered by their intrinsic flexibility. Second, the relationship between constitution and the so-called 'bioactive conformation' is far from obvious. Ideally, a direct structural study of the complex between a peptide and its receptor should answer both questions, but such a study is not possible, because opioid receptors are large membrane proteins, difficult to study by standard structural techniques. Thus, conformational studies of opioid peptides are still important for drug design and also for indirect receptor mapping. This review deals with conformational studies of natural opioid peptides in several solvents that mimic in part the different environments in which the peptides exert their action. None of the structural investigations yields a convincing bioactive conformation, but the global conformation of longer peptides in biomimetic environments can shed light on the interaction with receptors. Received 15 April 2001; received after revision 10 May 2001; accepted 11 May 2001     

Acute toxicity of folic acid in mice

The toxicity of folic acid (PGA) was studied in different inbred strains of mice. LD50 values of PGA by the i.p. route showed a unique toxicity pattern. In some strains, convulsions, ataxia and weakness were observed. Histopathological study in strains S/RVCri, BDF1, DBA/2 and DBA/2fNCri showed acute renal tubular necrosis.     

Persistent inverse maternal effect on corticosterone production in vitro

Adrenal cells from C57BL/Tb mice produced more steroid than those from DBA/2J mice. Reciprocal differences between both backcross and F1 hybrids showed a persistent maternal effect. Mothers with high output produce offspring with reduced hormone production when adult. Corticosterone output thus depends on maternal phenotype as well as on the genotype of the isolated cells.     

Genetic differences between inbred strains of mice; a new source of variation in high-sulphur keratins

Summary Proteins were extracted by improved techniques from the hair of inbred strains of Peru, C57BL/6 and CBA/Ca mice. The extracts were characterized by amino acid analysis and high resolution polyacrylamide gel electrophoresis, and previously unreported strain differences were observed. Genetic analysis confirmed them.We are grateful to Dr Speakman, Dr J. Findlay and Prof. J. Shire for helpful discussion, the Joint Sequencing Unit (Dept of Biochemistry/Genetics) for carrying out the amino acid analyses, and the mouse house staff. We also thank the S.R.C. for the grant GR/B/62877 which supported this work.     

Acute toxicity of folic acid in mice

Summary The toxicity of folic acid (PGA) was studied in different inbred strains of mice. LD₅₀ values of PGA by the i.p. route showed a unique toxicity pattern. In some strains, convulsions, ataxia and weakness were observed. Histopathological study in strains S/RVCri, BDF₁, DBA/2 and DBA/2fNCri showed acute renal tubular necrosis.Acknowledgment. The authors thank Ms N. Hasgekar for her technical assistance     

Production of passive cutaneous anaphylaxis (PCA) and reversed PCA by rat IgE antibody in the mouse

Summary Although IgE antibody is generally characterized as a homocytotropic antibody, it has been well known for some time that mouse IgE antibody causes potent sensitization of rat skin for PCA. The present study clearly shows, the reciprocal cross-sensitization of mouse skin with rat IgE molecules. PCA and RPCA were produced by rat IgE antibody in an inbred mouse strain, DS/Shi, though not in C3H/HeShi, C57BL/6JShi and BALB/cCrj strains. Sensitization of DS/Shi mouse skin for PCA with rat IgE antibody was comparable in sensitivity with that of rat skin, but lasted only for a short term in comparison with the long persistence in rat skin.     

Persistent inverse maternal effect on corticosterone production in vitro

Summary Adrenal cells from C57BL/Tb mice produced more steroid than those from DBA/2J mice. Reciprocal differences between both backcross and F1 hybrids showed a persistent maternal effect. Mothers with high output produce offspring with reduced hormone production when adult. Corticosterone output thus depends on maternal phenotype as well as on the genotype of the isolated cells.We thank Margaret MacKendrick for her help and the Medical Research Council for a research studentship for PRWW.     

The histidine triad nucleotide-binding protein 1 supports mu-opioid receptor–glutamate NMDA receptor cross-regulation

A series of pharmacological and physiological studies have demonstrated the functional cross-regulation between MOR and NMDAR. These receptors coexist at postsynaptic sites in midbrain periaqueductal grey (PAG) neurons, an area implicated in the analgesic effects of opioids like morphine. In this study, we found that the MOR-associated histidine triad nucleotide-binding protein 1 (HINT1) is essential for maintaining the connection between the NMDAR and MOR. Morphine-induced analgesic tolerance is prevented and even rescued by inhibiting PKC or by antagonizing NMDAR. However, in the absence of HINT1, the MOR becomes supersensitive to morphine before suffering a profound and lasting desensitization that is refractory to PKC inhibition or NMDAR antagonism. Thus, HINT1 emerges as a key protein that is critical for sustaining NMDAR-mediated regulation of MOR signaling strength. Thus, HINT1 deficiency may contribute to opioid-intractable pain syndromes by causing long-term MOR desensitization via mechanisms independent of NMDAR.     

Naloxone prevents the analgesic action of alpha-MSH in mice

alpha-MSH (0.1, 1, 10 micrograms) was administered intracerebroventricularly and its action on pain sensitivity was investigated by the hot-plate method in mice. alpha-MSH produced dose-dependent analgesia and this analgesic effect was prevented by naloxone (1 mg/kg, s.c.). It is possible that alpha-MSH may play a role in the mechanism of pain through endogeneous opioid systems.     

Production of passive cutaneous anaphylaxis (PCA) and reversed PCA by rat IgE antibody in the mouse

Although IgE antibody is generally characterized as a homocytotropic antibody, it has been well known for some time that mouse IgE antibody causes potent sensitization of rat skin for PCA. The present study clearly shows the reciprocal cross-sensitization of mouse skin with rat IgE molecules. PCA and RPCA were produced by rat IgE antibody in an inbred mouse strain. DS/Shi, though not in C3H/HeShi, C57BL/6JShi and BALB/cCrj strains. Sensitization of DS/Shi mouse skin for PCA with rat IgE antibody was comparable in sensitivity with that of rat skin, but lasted only for a short term in comparison with the long persistence in rat skin.     

Cytotoxicity of mouse peritoneal cells after alloimmunization]

CBA Mice were immunized by two intraperitoneal injections of 30 X 10(6) DBA/2 or C57BL/6 spleen cells at days--12 and--2. Peritoneal cell population was obtained at day zero by washing the peritoneal cavity of Mice. Adherent cells were then separated using a 2 hrs. incubation in "Falcon" plates followed by washing. This macrophage-rich peritoneal cell population was found nonspecifically cytotoxic against 51Cr labeled tumoral target cells: P815 X DBA/2 mastocytoma cells, EL4 X C57BL/L lymphoma cells and spontaneous lymphoma AKR cells (same H--2k as CBA). This adherent peritoneal cell cytoxicity was demonstrated after 24 hrs. incubation with the target cells. It was found in nonspecific combination as well as when using target cells syngeneic to the donor. These findings suggest that adherent peritoneal cell cytotoxicity could be at least partly due to macrophages and result from factor (s) released by sensitized lymphocytes in vivo in the same way as has been previously demonstrated in vitro.     

Expression of an antigen associated with Gross virus on the surfaces of murine cells producing an oncornavirus from the radioleukemia of C57BL/6 mice]

A leukemogenic viral complex was demonstrated in cultures of 13-3 C cell line derived from a C57BL/6, radiation leukemia virus (RadLV-Rs) induced tumor. Both 13-3C and leukemic cells induced in C57BL/6 mice by 13-3C virus carry a cell surface antigen associated with Gross leukemia virus (GCSAa). These findings point to a close similarity between these antigens and those of murine endogenous ecotropic viruses.     

Changes in brain dopamine levels and aggressive behavior with aging in 2 mouse strains

The genetic programming of brain monoamine changes with aging show remarkable differences in 2 mouse strains. A marked increase in dopamine occurred in 32-week-old grouped ICR mice and the males showed intense irritability and aggressive behavior. Brain amines changed only slightly in old C57BL6J mice and behavior remained benign. Old females showed similar amine changes but aggresive behavior did not occur in either strain.     

Naloxone prevents the analgesic action of α-MSH in mice

Summary -MSH (0.1, 1, 10 g) was administered intracerebroventricularly and its action on pain sensitivity was investigated by the hot-plate method in mice. -MSH produced dose-dependent analgesia and this analgesic effect was prevented by naloxone (1 mg/kg, s.c.). It is possible that -MSH may play a role in the mechanism of pain through endogeneous opioid systems.