Opioid receptors in the chicken brain

The term “opioid” generally refers to any substance, endogenous or synthetic peptide or non-peptide, which acts through any of the four major types of opioid receptors present in the cell membrane. Its effects can be stereospecifically blocked by pure narcotic antagonists like naloxone. The four major classes of endogenous opioid peptides, b-endorphin, enkephalin, dynorphin and nociceptin have been described as potential ligands for the m, d, k and opioid receptor like (ORL1) receptor binding sites. Opioid peptides and their receptors are implicated in the modulation of a number of behavioral and physiological functions in birds (e.g., reproduction, endocrinology, water balance, social behavior and painful stimulus) in a manner similar to that described for mammals. Opioids are (neuro) peptides, which are synthesized in the central nervous system (CNS) in the whole animal kingdom. They play a role both as neurotransmitters and neuromodulators.Opioid peptides and their receptors are implicated in the modulation of a number of behavioral and physiological functions in birds (e.g., reproduction, endocrinology, water balance, social behavior and painful stimulus) in a manner similar to that described for mammals. Nowadays, more attention is being paid to the effect of the opioid system on the water balance in birds. Opioid drugs were also introduced in veterinary medicine for pain relieving and anesthesia of birds. The studies reported in this dissertation were undertaken to further elucidate opioid receptors, the physiological intervention on receptor binding properties and their regulation. To this end, we evaluated the receptor binding profiles - the dissociation constant (Kd) and maximum binding capacity (Bmax) - in various brain regions. Following questions were addressed using the chicken as model. Are opioid binding profiles gender-, and/or age-dependent in the chicken brain? Are opioid binding profiles affected by dehydration in the chicken brain? Are opioid binding profiles testosterone-dependent?, and Are opioid binding profiles affected by opioid antagonist? To achieve this, the opioid binding sites were determined in six brain regions - frontal cortex, lateral septum, striatum, amygdala, hippocampus and hypothalamus - of chicken using [3H]diprenorphine ([3H]DPN) as the radioligand. Three age groups of chickens, 10-day-, 10-week- and 26-38-week-old, were used. Following situations were considered (a) the effects of hydration (b) the effect of testosterone treatments (performed in castrated male chickens) and (c) the effect of opioid antagonist, naltrexone. Experiment I, opioid binding sites were investigated in 10-day-, 10-week- and 26-38-week-old female and male chickens. The linear Scatchard plot of [3H]DPN binding in all experiments indicated a single binding site with high affinity. The mean of Bmax was as following: 0.27-0.44 (female), 0.23-0.33 (male); 0.21-0.48 (female), 0.21-0.41 (male); 0.19-0.34 (female), 0.17-0.34 (male) pmol/mg in 10-day-, 10-week- and 26-38-week-old chickens, respectively. The mean of Kd was as follow:  1.13-1.80 (female), 0.72-1.01 (male); 0.28-0.48 (female), 0.27-0.83 (male); 0.33-0.75 (female), 0.26-0.58 (male) nM in 10-day-, 10-week- and 26-38-week-old chickens, respectively. The highest concentration of opioid binding sites was found in the frontal cortex and the lowest concentration in the hypothalamus both in males and females. The Bmax of opioid binding sites decreased significantly from 10-day-old to adulthood in the frontal cortex, striatum and amygdala regions. While there was no such a decrease in males. The data also indicated that the sex related change in opioid binding reflects a change in the Kd in males more than in females. Experiment II, groups of 10-day-, 10-week- and 26-38-week-old chickens were deprived from water for 1 or 2 days. Dehydration induced an increase of plasma AVT levels and had some impacts on the characteristics of opioid binding sites in the brain.  Most of the changes of opioid binding site were in young female chickens. The results also indicated that dehydration induced both decrease (frontal cortex, striatum and amygdala: 10-day-old females; lateral septum: adult females; and hypothalamus: 10-day-old males) and increase (lateral septum, hippocampus and hypothalamus: 10-week-old females) of Bmax in chicken brains. Such changes were accompanied with alterations in receptor affinity. Experiment III, male chickens were castrated and sham-operated for 7 months and were treated with testosterone proprionate or testosterone proprionate and naltrexone. The Bmax value in the brain of castrated chickens was higher than that found in intact animals. This increase was testosterone-dependent, since testosterone substitution reduced the binding capacity to the control levels. The extent of opioid binding site changes after naltrexone administration was more pronounced in the Kd than in the Bmax values. The regional differences in naltrexone-induce changes imply that the drug does not have uniform effects throughout the brain. The Bmax was significantly increased in the hypothalamus only. In conclusion, these results demonstrate, that like in some mammals, opioid binding sites are involved in modulation of water balance and reproduction in chickens. Such modulations are age-, sex- and brain region dependent.