Clare H. Spicer

Early life programming of fear conditioning and extinction in adult male rats

The early rearing environment programs corticolimbic function and neuroendocrine stress reactivity in adulthood. Although early environmental programming of innate fear has been previously examined, its impact on fear learning and memory later in life remains poorly understood. Here we examined the role of the early rearing environment in programming fear conditioning and extinction in adult male rats. Pups were subjected to maternal separation (MS; 360 min), brief handling (H; 15 min), or animal facility rearing (AFR) on post-natal days 2-14. As adults, animals were tested in a 3-day fear learning and memory paradigm which assessed the acquisition, expression and extinction of fear conditioning to an auditory cue; the recall of extinction was also assessed. In addition, contextual fear was assessed prior to cued extinction and its recall. We found that the acquisition of fear conditioning to the cue was modestly impaired by MS. However, no early rearing group differences were observed in cue-induced fear expression. In contrast, both the rate of extinction and extinction recall were attenuated by H. Finally, although contextual fear was reduced after extinction to the cue, no differences in context-induced fear were observed between the early rearing groups. These results add to a growing body of evidence supporting an important role for early environmental programming of fear conditioning and extinction. They also indicate that different early rearing conditions can program varying effects on distinct fear learning and memory processes in adulthood.

Early life programming of innate fear and fear learning in adult female rats

The early rearing environment can impact on emotional reactivity and learning later in life. In this study the effects of neonatal maternal separation (MS) on innate fear and fear learning were assessed in the adult female rat. Pups were subjected to MS (360 min), brief handling (H; 15 min), or animal facility rearing (AFR) on post-natal days 2-14. In the first experiment, innate fear was tested in the open field. No differences between the early rearing groups were observed in unconditioned fear. In the second experiment, separate cohorts were used in a 3-day fear learning paradigm which tested the acquisition (Day 1), expression and extinction (both Day 2) of conditioning to an auditory cue; extinction recall was determined as well (Day 3). Contextual fear conditioning was also assessed prior to cue presentations on Days 2 and 3. Whereas MS attenuated the acquisition and expression of fear conditioning to the cue, H potentiated extinction learning. Cue-induced fear was reduced on Day 3, compared to Day 2, indicating that the recall of extinction learning was evident; however, no early rearing group differences in extinction recall were observed. Similarly, while contextual fear was decreased on Day 3, compared to Day 2, there were no differences between the early rearing groups on either day tested. The present findings of altered cue-conditioned fear learning, in the absence of innate fear changes, lend further support for the important role of the early rearing environment in mediating cognition in adulthood.

Effects of chronic infusion of neurotensin and a neurotensin NT1 selective analogue PD149163 on amphetamine-induced hyperlocomotion

Neurotensin (NT) has been proposed as an endogenous antipsychotic based in part on the similarity in behavioural effects to antipsychotic drugs, for example, attenuation of both amphetamine-induced hyperlocomotion (AH) and amphetamine disrupted pre-pulse inhibition in the rat. However, there is some evidence that repeated administration of NT or an analogue produces behavioural tolerance to such effects. The present experiments sought to confirm and extend these findings by testing the effects on AH of 7 days central administration of NT and the NT 1 selective analogue PD 149163 and the effects of 21 days central administration of NT. NT and PD149163 continuously administered for 7 days produced no effect on AH (in contrast to attenuation with a single injection here and previously reported), whereas 21 days of NT administration potentiated AH. Together, these studies report that the effects of NT or a NT analogue on AH depends on the duration of administration of peptide. The results are discussed in comparison with the reported antipsychotic properties of acute administration of NT and possible mechanisms involving NT1 receptors.

Contribution of serotonin and dopamine to changes in core body temperature and locomotor activity in rats following repeated administration of mephedrone.

The psychoactive effects of mephedrone are commonly compared with those of 3,4‐methylenedioxymethamphetamine, but because of a shorter duration of action, users often employ repeated administration to maintain its psychoactive effects. This study examined the effects of repeated mephedrone administration on locomotor activity, body temperature and striatal dopamine and 5‐hydroxytryptamine (5‐HT) levels and the role of dopaminergic and serotonergic neurons in these responses. Adult male Lister hooded rats received three injections of vehicle (1 ml/kg, i.p.) or mephedrone HCl (10 mg/kg) at 2 h intervals for radiotelemetry (temperature and activity) or microdialysis (dopamine and 5‐HT) measurements. Intracerebroventricular pre‐treatment (21 to 28 days earlier) with 5,7‐dihydroxytryptamine (150 µg) or 6‐hydroxydopamine (300 µg) was used to examine the impact of 5‐HT or dopamine depletion on mephedrone‐induced changes in temperature and activity. A final study examined the influence of i.p. pre‐treatment (−30 min) with the 5‐HT1A receptor antagonist WAY‐100635 (0.5 mg/kg), 5‐HT1B receptor antagonist GR 127935 (3 mg/kg) or the 5‐HT7 receptor antagonist SB‐258719 (10 mg/kg) on mephedrone‐induced changes in locomotor activity and rectal temperature. Mephedrone caused rapid‐onset hyperactivity, hypothermia (attenuated on repeat dosing) and increased striatal dopamine and 5‐HT release following each injection. Mephedrone‐induced hyperactivity was attenuated by 5‐HT depletion and 5‐HT1B receptor antagonism, whereas the hypothermia was completely abolished by 5‐HT depletion and lessened by 5‐HT1A receptor antagonism. These findings suggest that stimulation of central 5‐HT release and/or inhibition of 5‐HT reuptake play a pivotal role in both the hyperlocomotor and hypothermic effects of mephedrone, which are mediated in part via 5‐HT1B and 5‐HT1A receptors.

Basolateral amygdala activity during the retrieval of associative learning under anesthesia.

Associative learning can occur under anesthesia and its neural correlates have begun to be elucidated. During discrimination learning under anesthesia in rats, lateral amygdala excitability increases in response to a conditioned stimulus (CS+) previously paired with electrical stimulation of the paw but not to another stimulus presented alone (CS-). Similarly, medial prefrontal cortex activity increases selectively during CS+ presentation after discrimination learning but this occurs only in neurons receiving input from the basolateral amygdala (BLA), the main source of amygdaloid projections to this region. However, BLA activity during discrimination learning under anesthesia has not been investigated. Here we used in vivo electrophysiology to examine BLA activity before and after associative learning and during later memory retrieval in anesthetized rats. We examined extracellular unit and local field potential (LFP) activity using an auditory discrimination learning paradigm. Rats were repeatedly presented with two distinct sounds, one of which was paired with electrical stimulation of the paw. One hour later, the paired sound (CS+) was presented alone along with the sound not paired with electrical stimulation (CS-). We found increased unit firing late (1 h) but not early (5 min) after learning. LFP power was increased both early and late after learning. In control experiments we also found increased unit and LFP activity late after electrical stimulation alone. After discrimination learning, unit firing increased in response to CS+, but not CS-, presentation. LFP power also showed a modest increase during CS+, compared to CS-, presentation. These findings suggest that discrimination learning under anesthesia can occur at the neural level in BLA. The potential relevance of these results is discussed in relation to previous studies examining neural activity during fear learning and memory processing in conscious animals.

Neural correlates of hyperalgesia in the monosodium iodoacetate model of osteoarthritis pain

Background The mechanisms driving osteoarthritic pain remain poorly understood, but there is increasing evidence for a role of the central nervous system in the chronification of pain. We used functional magnetic resonance imaging to investigate the influence of a model of unilateral knee osteoarthritis on nociceptive processing. Results Four to five weeks post intra-articular injection of monosodium iodoacetate (MIA, 1 mg) into the left knee, Sprague Dawley rats were anesthetized for functional magnetic resonance imaging studies to characterize the neural response to a noxious stimulus (intra-articular capsaicin injection). In a two-arm cross-over design, 5 µM/50 µl capsaicin was injected into either the left knee (n = 8, CAPS-MIA) or right control knee (n = 8, CAPS-CON), preceded by contralateral vehicle (SAL) injection. To assess neural correlates of mechanical hyperalgesia, hindpaws were stimulated with von Frey hairs (8 g: MIA; 15 g: control knee, based on behavioral withdrawal responses). The CAPS-MIA group exhibited significant activation of the periaqueductal gray, unilateral thalamus and bilateral mensencephalon, superior-colliculus, and hippocampus, with no significant activation in the other groups/conditions. Capsaicin injection increased functional connectivity in the mid-brain network and mediodorsal thalamic nucleus, hippocampus, and globus pallidus, which was significantly stronger in CAPS-MIA compared to CAPS-CON groups. Mechanical stimulation of the hyperalgesic (ipsilateral to MIA knee) and normalgesic (contralateral) hindpaws evoked qualitatively different brain activation with more widespread brainstem and anterior cingulate (ACC) activation when stimulating the hyperalgesic paw, and clearer frontal sensory activation from the normalgesic paw. Conclusions We provide evidence for modulation of nociceptive processing in a chronic knee osteoarthritis pain model with stronger brain activation and alteration of brain networks induced by the pro-nociceptive stimulus. We also report a shift to a medial pain activation pattern following stimulation of the hyperalgesic hindpaw. Taken together, our data support altered neural pain processing as a result of peripheral and central pain sensitization in this model.

Systemic administration of WY-14643, a selective synthetic agonist of peroxisome proliferator activator receptor-alpha, alters spinal neuronal firing in a rodent model of neuropathic pain

Abstract Background and aims The clinical management of chronic neuropathic pain remains a global health challenge. Current treatments are either ineffective, or associated with unwanted side-effects. The development of effective, safe therapies requires the identification of novel therapeutic targets using clinically relevant animal models of neuropathic pain. Peroxisome proliferator activated receptor alpha (PPARα), is a member of the nuclear hormone family of transcription factors, which is widely distributed in the peripheral and central nervous systems. Pharmacological studies report antinociceptive effects of PPARα agonists following systemic administration in rodent models of neuropathic pain, however the neuronal mechanisms and sites of action mediating these effects are unclear. The aim of this study was to investigate the effects of systemic administration of the synthetic PPARα agonist, WY-14643 on mechanically-evoked responses of spinal cord dorsal horn wide dynamic range (WDR) neurones in the spinal nerve ligated (SNL) model of neuropathic pain in rats. In addition, comparative molecular analysis of mRNA coding for PPARα and PPARα protein expression in the spinal cord of sham-operated and neuropathic rats was performed. Methods Lumbar L5–L6 spinal nerve ligation was performed in male Sprague–Dawley rats (110–130 g) under isoflurane anaesthesia. Sham controls underwent similar surgical conditions, but without ligation of the L5–L6 spinal nerves. Hindpaw withdrawal thresholds were measured on the day of surgery -day 0, and on days- 2, 4, 7, 10 and 14 post-surgery. At day 14 extracellular single-unit recordings of spinal (WDR) dorsal horn neurons were performed in both sham and SNL neuropathic rats under anaesthesia. The effects of intraperitoneal (i.p.) administration of WY-14643 (15 and 30 mg/kg) or vehicle on evoked responses of WDR neurons to punctate mechanical stimulation of the peripheral receptive field of varying bending force (8–60 g) were recorded. In a separate cohort of SNL and sham neuropathic rats, the expression of mRNA coding for PPARα and protein expression in the ipsilateral and contralateral spinal cord was determined using quantitative real time polymerase chain reaction (qRT-PCR) and western blotting techniques respectively. Results WY-14643 (15 and 30mg/kg i.p.) rapidly attenuated mechanically evoked (8, 10 and 15g) responses of spinal WDR neurones in SNL, but not sham-operated rats. Molecular analysis revealed significantly increased PPARα protein, but not mRNA, expression in the ipsilateral spinal cord of SNL, compared to the contralateral side in SNL rats. There were no changes in PPARα mRNA or protein expression in the sham controls. Conclusion The observation that levels of PPARα protein were increased in ipsilateral spinal cord of neuropathic rats supports a contribution of spinal sites of action mediating the effects of systemic WY-14643. Our data suggests that the inhibitory effects of a PPARα agonist on spinal neuronal responses may account, at least in part, for their analgesic effects of in neuropathic pain. Implication Selective activation of PPARα in the spinal cord may be therapeutically relevant for the treatment of neuropathic pain.

Neonatal maternal separation alters reward-related ultrasonic vocalizations in rat dams.

We examined the effects of brief or long durations of repeated maternal separation (MS) on ultrasonic vocalizations (USVs) in rat dams. 50-kHz USVs putatively identified as maternal in origin were emitted only immediately after pups were returned following MS. Maternal USV emission also depended on separation duration and pup age. Given that 50-kHz USVs are emitted in response to rewarding stimuli, MS may induce duration-dependent effects on maternal positive affect.