As described above, opioid dependence is associated with a characteristic withdrawal syndrome that appears with the abrupt termination of opioid administration or administration of competitive opioid receptor antagonists, such as naloxone (termed “precipitated withdrawal”). The neural substrates for the physical and affective signs of opioid withdrawal can be dissociated. In rats, physical dependence has been characterized by an abstinence syndrome that includes the appearance of ptosis, teeth chattering, wet dog shakes (which are exactly what the phrase suggests), and diarrhea. This syndrome can be precipitated in dependent animals by systemic injections of opioid antagonists. Motivational withdrawal includes changes in brain reward thresholds, the disruption of trained operant behavior for food reward, and conditioned place aversions. Precipitated opioid withdrawal is associated with increased brain reward thresholds at doses of naloxone that do not elicit physical signs of withdrawal, which, as mentioned above, indicates that the mechanisms of the physical and motivational components of opioid withdrawal are different (Figure 5.13).

Opioid dependence and tolerance can begin with a single administration of the drug. Single opioid injections can induce a state of acute opioid dependence in both humans and animals. Acute dependence is particularly evident when evaluating motivational measures of withdrawal, such as the suppression of operant responding for food, place aversion, and brain stimulation reward (Figure 5.14). Repeated morphine administration results in a progressive increase in the potency of naloxone to elicit precipitated withdrawal. Greater shifts in potency were observed when naloxone was administered on all treatment days but only in the testing situation and not in the home cage where the rat had received no drug injections, suggesting an important conditioning component of acute dependence. An hypothetical, parallel human example is that of an individual with opioid use disorder who undergoes withdrawal in a particular environment and experiences conditioned withdrawal when exposed again to that environment. A lasting association between a specific environment and the aversive stimulus effects of opioid withdrawal can persist for a long time after the acute withdrawal state has dissipated.

FIGURE 5.13 Naloxone dose-dependently elevated intracranial self-stimulation reward thresholds in rats (reflecting a decrease in reward function) in the morphine group but not in the placebo group. The minimum dose of naloxone that elevated ICSS thresholds in the morphine group was 0.01 mg/kg (∗p < 0.05, compared with vehicle). These results show that low doses of naloxone can “precipitate” a dysphoric-like effect (increased reward thresholds) in morphine-dependent rats at doses that do not cause any change in nondependent control rats. [Taken with permission from Schulteis G, Markou A, Gold LH, Stinus L, Koob GF. Relative sensitivity to naloxone of multiple indices of opiate withdrawal: a quantitative dose-response analysis. Journal of Pharmacology and Experimental Therapeutics, 1994, (271), 1391–1398.]

Animal studies have observed hyperalgesia during spontaneous (in which drug administration is simply ceased) or precipitated (in which an antagonist is administered) opioid withdrawal following acute or chronic opioid exposure (for further reading, see Simonnet and Rivat, 2003). Some studies have even suggested that pain sensitization might develop while the subjects are still exposed to the opioid. Moreover, heroin effectively facilitated pain in rats after just one exposure to the drug (Figure 5.15). This phenomenon may indicate the actual sensitization of pain systems because both the magnitude and duration of hyperalgesia increased as a function of opioid administration. For example, the first heroin administration may induce moderate hyperalgesia for 2 days, whereas the fifth injection of the same dose may induce hyperalgesia for 6 days. When the opioid is administered repeatedly, for example once daily for 2 weeks, a gradual and dose-dependent decrease in nociceptive threshold is observed that lasts for several weeks after drug administration (Figure 5.15). A small dose of heroin that was otherwise ineffective at triggering delayed hyperalgesia in non-heroin-treated rats enhanced pain sensitivity for several days after a series of heroin injections, suggesting the occurrence of pain sensitization. The effectiveness of naloxone in precipitating hyperalgesia in rats that had recovered their pre-drug nociceptive thresholds after single or repeated heroin injections indicates that heroin-deprived rats were in a new biological state associated with a functional balance between opioid-dependent analgesic systems and pronociceptive systems. Such a finding may also be another explanation of tolerance, in which a prolonged increase occurs in the excitability of nociceptive pathways that increasingly opposes analgesia induced by repeated opioid administration. Thus, a neuronal memory, characterized by a vulnerable state, may remain long after the complete wash-out of the drug and when apparent equilibrium near the pre-drug state has been re-established. These long-term effects may include the expression of other allostatic changes, such as negative emotional states, long after withdrawal from the drug.

FIGURE 5.14 Mean time spent by rats in each of three compartments of a place conditioning apparatus. All morphine/naloxone treatment conditions represent two conditioning cycles, except where noted. As time spent in the naloxone-paired compartment declines as a function of naloxone dose, this extra time appears to be randomly spent in either the vehicle-paired or neutral compartment. These data show that low doses of naloxone can “precipitate” an aversive-like effect in rats, even following an acute administration of morphine, suggesting that the opponent process (b process) produced by opioids begins with the first opioid injection. [Taken with permission from Azar MR, Jones BC, Schulteis G. Conditioned place aversion is a highly sensitive index of acute opioid dependence and withdrawal. Psychopharmacology, 2003, (170), 42–50.]

The mechanism for such hyperalgesia may involve activation of glutamatergic systems. A noncompetitive glutamate receptor antagonist prevented both the long-lasting heroin-induced enhancement in pain sensitivity and naloxone-precipitated hyperalgesia. A parallel has been found between the development of thermal hyperalgesia in rats and tolerance to the analgesic effects of morphine administered for eight consecutive days. The concurrent development of tolerance was prevented by co-administration of morphine with a glutamate receptor antagonist and protein kinase C inhibitor.

FIGURE 5.15 Effects of heroin on vocalization threshold to paw pressure in rats. (A) Rats received a subcutaneous injection of 2.5 mg/kg heroin or saline after the determination of basal paw pressure-inducing vocalization, and then the values were determined every 30 min for 6 h. Each point represents the mean paw pressure ± SEM of 8–10 animals. ∗p < 0.05, ∗∗p < 0.01. These data show an opponent process-like effect on pain thresholds following a single dose of heroin in the rat. Initially, for 3 hours, heroin produced analgesia (an increase in pain thresholds). However, after the analgesia disappeared, hyperalgesia (a decrease in pain thresholds) developed (see text for a definition of hyperalgesia). [Taken with permission from Laulin JP, Larcher A, Celerier E, Le Moal M, Simonnet G. Long-lasting increased pain sensitivity in rat following exposure to heroin for the first time. European Journal of Neuroscience, 1998, (10), 782–785.]

Studies of the neurobiological substrates of physical dependence have revealed multiple sites of action, including the periaqueductal gray, dorsal thalamus, and locus coeruleus. However, the brain sites responsible for the motivational and emotional changes associated with opioid withdrawal have been localized to the nucleus accumbens and amygdala. The aversive stimulus effects of opioid withdrawal that form the basis for its motivational effects can readily be measured in rats using conditioned place aversion or the suppression of operant responding for a food reward. Thus, during chronic morphine exposure, neural elements in the nucleus accumbens and amygdala may become sensitized to opioid antagonists and may be partially responsible for the negative stimulus effects of opioid withdrawal (Figure 5.16).

FIGURE 5.16 The effect of intracerebral methylnaloxonium paired with a particular environment on the amount of time spent by rats in that environment during an injection-free test session. Values represent the median difference between the postconditioning score and the preconditioning score. Darkened bars represent those doses where the conditioning scores were significantly different from the preconditioniong scores. Significance was set at p < 0.02 to control for multiple comparisons. These results show that a naloxone derivative (methylnaloxium, an opioid antagonist) when injected directly into the brain can produce an aversive-like effect in opioid-dependent animals, with the lowest doses producing an aversive effect, particularly in the nucleus accumbens and central nucleus of the amygdala. [Taken with permission from Stinus L, Le Moal M, Koob GF. Nucleus accumbens and amygdala are possible substrates for the aversive stimulus effects of opiate withdrawal. Neuroscience, 1990, (37), 767–773.]

FIGURE 5.17 Unlimited daily access to heroin escalated heroin intake and decreased the excitability of brain reward systems. (Left) Heroin intake (20 μg per infusion) in rats during limited (1 h) or unlimited (23 h) self-administration sessions. ∗∗∗p < 0.001, effect of access (1 or 23 h). (Right) Percentage change from baseline ICSS thresholds in 23 h rats. Reward thresholds, assessed immediately after each daily 23 h self-administration session, became progressively more elevated as exposure to self-administered heroin increased across sessions. ∗p < 0.05, effect of heroin on reward thresholds. These data show that the overall increase in thresholds over days associated with extended access to heroin paralleled the overall increase in drug taking over days (termed escalation) associated with extended access. [Taken with permission from Kenny PJ, Chen SA, Kitamura O, Markou A, Koob GF. Conditioned withdrawal drives heroin consumption and decreases reward sensitivity. Journal of Neuroscience, 2006, (26), 5894–5900.]

Data from conditioned place aversion studies and unlimited access to intravenous heroin self-administration provide a neuropharmacological framework for the neurocircuitry associated with opioid addiction. Studies of lesions of the extended amygdala while measuring precipitated withdrawal-induced place aversions support a role for the extended amygdala in the aversive stimulus effects of opioid withdrawal. Lesions of the central nucleus of the amygdala, one part of the extended amygdala, block the development of morphine withdrawal-induced place aversion but have less of an effect on the somatic signs of withdrawal.

Neurochemical elements within the extended amygdala involved in the aversive stimulus effects of opioid withdrawal include corticotropin-releasing factor (CRF) and norepinephrine. Blockade of CRF receptors in the central nucleus of the amygdala blocked conditioned place aversion produced by opioid withdrawal. Inactivation of noradrenergic (norepinephrine) function in the bed nucleus of the stria terminalis, another region of the extended amygdala, blocked the aversive motivational effects of opioid withdrawal. Injections of a β-adrenergic receptor antagonist or α2 receptor agonist into the lateral bed nucleus of the stria terminalis blocked precipitated opioid withdrawal-induced place aversion. Norepinephrine concentrations in the bed nucleus of the stria terminalis represent 10% of the brain’s norepinephrine. Blockade of norepinephrine receptors in the central nucleus of the amygdala with β-adrenergic antagonists attenuated morphine withdrawal-induced conditioned place aversion.

Using the intravenous self-administration model of extended access, rats will steadily increase heroin intake, showing a pattern of responding not unlike humans with opioid addiction (Figure 5.17). The daily pattern of intake shifts, with decreases in food and water intake and the development of motivational signs of dependence (for further reading, see Chen et al., 2006). The same neuropharmacological systems implicated in the negative emotional effects of opioid withdrawal have been shown to be important for the compulsive drug taking and seeking associated with extended-access intravenous self-administration in animal models. Both CRF antagonists and α1 noradrenergic antagonists dose-dependently decrease the compulsive drug intake observed in opioid-dependent rats.

Neuronal plasticity occurs in the nucleus accumbens and amygdala with the development of morphine dependence. Dopamine neuron firing and extracellular dopamine levels are decreased during opioid withdrawal. Chronic morphine administration is also associated with a decrease in the size of ventral tegmental area dopamine neurons and an increase in the sensitivity to dopamine antagonists. The cellular changes that occur during opioid withdrawal are accompanied by increased GABA activity and increased metabotropic glutamate receptor sensitivity, which both decrease glutamate release in the ventral tegmental area and lead to decreased dopamine cell firing. Chronic opioid administration, therefore, has multiple effects on the neuropharmacological systems that interface with the ventral tegmental area and extended amygdala, which may provide the cellular basis for the neuroadaptations associated with the development of dependence. Opioids act indirectly via GABA and glutamate systems to activate reward pathways, and these same systems show neuroadaptations with chronic opioid exposure.

Acute opioid receptor activation leads to the inhibition of adenylyl cyclase (Figure 5.18), the cyclic adenosine monophosphate (cAMP) protein phosphorylation cascade via the recruitment of Gi and related G proteins, activation of voltage-gated Ca2+ channels, and activation of K+ channels. More K+ flows out of the cell, and less Ca2+ flows into the cell. Reductions in cellular Ca2+ levels alter Ca2+-dependent protein phosphorylation cascades. Alterations in the activity of these protein phosphorylation cascades, which can vary among different cell types, lead to the regulation of still other ion channels, which further contributes to the acute effects of the drug.

FIGURE 5.18 Upregulation of the cyclic adenosine monophosphate (cAMP) pathway as a mechanism of opioid tolerance and dependence. Opioids acutely inhibit the functional activity of the cAMP pathway (indicated by cellular levels of cAMP and cAMP-dependent protein phosphorylation). With continued opioid exposure, functional activity of the cAMP pathway gradually recovers and then increases far above control levels following removal of the opioid (e.g., by administration of the opioid receptor antagonist naloxone). These changes in the functional state of the cAMP pathway are mediated by the induction of adenylyl cyclase and protein kinase A in response to chronic administration of opioids. The induction of these enzymes accounts for the gradual recovery in the functional activity of the cAMP pathway that occurs during chronic opioid exposure (tolerance and dependence) and activation of the cAMP pathway observed upon removal of opioid (withdrawal). [Taken with permission from Nestler EJ. Historical review: molecular and cellular mechanisms of opiate and cocaine addiction. Trends in Pharmacological Sciences. 2004. (25), 210–218.]

Chronic exposure to opioids increases adenylyl cyclase, leading to other perturbations in intracellular messenger pathways, such as in protein phosphorylation mechanisms. These perturbations eventually elicit long-term adaptations that can lead to changes in many other neural processes within target neurons. These target neurons include those that trigger the long-term effects of the drugs, eventually leading to tolerance, dependence, withdrawal, sensitization, and addiction (Figure 5.18). Opioid withdrawal increases the expression of the transcription factor Fos, a marker of neural activation, with particular sensitivity in the extended amygdala. Such Fos changes correlate with the motivational effects (place aversion) rather than somatic effects of opioid withdrawal (for further reading, see Nestler, 2001).

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