Forebrain contribution to the induction of a brainstem correlate of conditioned taste aversion: II. Insular (gustatory) cortex

The induction of c-Fos-like immunoreactivity (c-FLI) in the intermediate division of the nucleus of the solitary tract (iNTS) has been shown to be a cellular correlate of the behavioral expression of a conditioned taste aversion (CTA). To further define neuroanatomical structures and pathways that contribute to this cellular response and to CTA learning in general, electrolytic lesions of insular (gustatory) cortex (IC) were combined with immunostaining for c-FLI. Rats were given either unilateral or bilateral electrolytic lesions of insular cortex or `sham' operations. Following surgery, `paired' animals were given a single conditioning trial consisting of intraoral infusion of 5-ml 0.15% sodium–saccharin followed by injection with LiCl (0.15 M, 20 ml/kg, i.p.) while `unpaired' controls received a non-contingent saccharin–LiCl presentation. Rats with bilateral lesions showed no behavioral evidence of having acquired a CTA. Increases in c-FLI in iNTS were evident, but reduced, relative to `sham' animals. Rats with unilateral-lesions displayed a CTA by rejecting the saccharin, although increases in c-FLI on the side of the iNTS ipsilateral to the lesion were reduced relative to that seen in `sham' animals. A comparison of these results with those obtained after amygdala lesions supports the conclusion that amygdala and insular cortex are necessary, but not sufficient, for the behavioral expression of a CTA.     

Insular cortex and amygdala lesions differentially affect acquisition on inhibitory avoidance and conditioned taste aversion

These experiments examined the effects of NMDA-induced lesions of the amygdala and insular (gustatory) cortex (IC) on inhibitory avoidance learning and conditioned taste aversion (CTA) in rats. IC lesions, but not amygdala lesions, disrupted CTA. In contrast, lesions of either brain region disrupted inhibitory avoidance learning. These findings support the view that the IC is strongly involved in the acquisition of external as well as visceral aversively motivated behavior. Despite extensive functional interconnections, these 2 brain regions appear to have different roles in mediating different forms of aversively based learning.     

C-fos expression in the rat brain following lithium chloride-induced illness

The present study examined c-Fos expression in selected brain areas consequent to administration of lithium chloride, the typical illness-inducing agent used in laboratory studies of conditioned taste aversion. The results replicated previous findings of significant c-Fos expression in the parabrachial nucleus, the central nucleus of the amygdala and the basolateral amygdala. New findings indicate significant lithium-induced c-Fos in the gustatory region of the thalamus and the bed nucleus of the stria terminalis but not in the insular cortex. The results are discussed with respect to the neural substrates of conditioned taste aversion.     

Induction of a brainstem correlate of conditioned taste aversion expression: role of the pontine parabrachial nucleus

Increases in Fos-like immunoreactivity (FLI) in the intermediate division of the nucleus of the solitary tract (iNTS) are seen following the expression of a conditioned taste aversion (CTA). In studies limited to behavioral assessment, the pontine parabrachial nucleus (PBN) has been demonstrated to play a critical role in the acquisition, but not the expression, of CTAs. To better define the role of the PBN in taste aversion learning, the present study examined the effects of PBN lesions on FLI in iNTS in animals with lesions placed either before or after CTA training. As is the case with behavioral expression of a CTA, timing of PBN lesions was found to be critical. Lesions placed prior to conditioning blocked evidence of conditioning, including both taste rejection and FLI in iNTS. Lesions placed after conditioning, but before testing, did not interfere with either taste rejection or FLI. These results support and extend prior claims that PBN is critical for CTA acquisition but not expression. They also demonstrate that input from PBN to iNTS is not necessary for the FLI seen there during CTA expression.     

Establishing aversive, but not safe, taste memories requires lateralized pontine-cortical connections

Aversive and safe taste memory processing is dramatically disrupted by bilateral lesions of the pontine parabrachial nucleus (PBN). To determine how such lesions affect patterns of neuronal activation in forebrain, lesions were combined with assessment of cFos-like immunoreactivity (FLI) in insular cortex (IC) and amygdala after conditioned taste aversion (CTA) training. Increases in FLI in amygdala and IC, which are normally seen following novel (versus familiar) CS-US pairing, were eliminated after PBN lesions. This suggests that PBN lesions prevent transmission of critical CS and US information to forebrain regions for the processing of both aversive and safe taste memories. Unilateral asymmetrical lesions of PBN and IC blocked CTA acquisition as well as normal patterns of FLI in amygdala after novel CS-US pairing, an effect not seen when unilateral lesions were confined to a single hemisphere. The crossed-disconnection experiments provide compelling evidence that functional interactions between PBN and IC are required for CTA acquisition, but not for safe taste memory formation and retrieval. The dissociation between effects of the different types of lesions on safe and aversive taste memories supports emerging evidence that the neural underpinnings of the two types of taste learning differ.     

Conditioned enhancement of antibody production is disrupted by insular cortex and amygdala but not hippocampal lesions.

Pavlovian conditioning procedures can be used to activate the immune system. A reliable conditioned increase of antibody production can be obtained in rats that have previously received a gustative or odor stimulus as the conditioned stimulus paired with an antigen, by exposing the animals to the conditioned stimulus alone. We showed evidence that an excitotoxic lesion bilaterally applied into the insular cortex or the amygdala, but not into the dorsal hippocampus, impaired the acquisition of both odor and gustatory conditioned immune enhancement. We found no effects of lesions on normal antibody production. These results suggest that the amygdala and the insular cortex are involved in the neural-immune interactions that mediate conditioned immunity.     

Differential involvement of cortical muscarinic and NMDA receptors in short- and long-term taste aversion memory

In conditioned taste aversion, an animal avoids a taste previously associated with toxic effects, and this aversive memory formation requires an intact insular cortex. In this paper, we investigated the possible differential involvement of cholinergic and glutamatergic receptors in the insular cortex in short-term memory (STM) and long-term memory (LTM) of taste aversion in rats. Taste aversion was induced by intraperitoneal administration of lithium chloride (a malaise-inducing drug) 15 min after experience with an unfamiliar taste. In order to test STM and LTM of taste aversion, taste stimulus was again presented 4 h and 72 h after lithium injection, respectively. During the acquisition, microinjection of the muscarinic antagonist, scopolamine, in the insular cortex before, but not after, the presentation of the new taste, abolished STM as well as LTM. Blockade of the NMDA receptor, in the insular cortex, by AP5 before, but not after, the presentation of the taste stimulus, impaired LTM but left STM intact. Moreover, when injected 1 h after malaise induction (i.e., during taste-illness association), AP5 disrupted both STM and LTM. These results suggest that activation of muscarinic receptors in the insular cortex is involved in the acquisition of taste memory, whereas NMDA receptors participate in taste memory consolidation. These data demonstrate that different neurochemical mechanisms subserve different memory phases. NMDA receptors are also probably involved in processing the visceral input, thus allowing subsequent taste-illness association. This indicates that in the same cortical area the same neurotransmitter system can be involved in distinct processes: taste memory consolidation vs. taste-illness association.     

Immune activation paired with intraoral sucrose conditions oral rejection

The effects of lipopolysaccharide (LPS) and LiCl on conditioned taste aversion acquisition using intraoral infusions as the method of taste delivery was examined. Rats received two pairings of an intraorally delivered sucrose (5 ml) taste with the effects of a systemic injection of LPS, LiCl or NaCl. The magnitude of conditioning was quantified by scoring taste reactivity responses to a brief intraoral infusion of sucrose in the absence of any drug injection. Rats previously conditioned with LiCl or LPS displayed clear evidence of conditioned aversion with increased oral rejection responses relative to saline controls. Our results suggest activation of the immune system with LPS can condition consummatory aspects of ingestion when this conditioning involves intraoral fluid presentation.     

cFos induction during conditioned taste aversion expression varies with aversion strength

Fos-like immunoreactivity (FLI) can indicate the location of neurons activated following expression of conditioned taste aversion (CTA). After one conditioning trial FLI has been identified in the intermediate nucleus of the solitary tract (iNTS) with little expression in other brain regions. The present study assessed the effect of increasing aversion strength on the magnitude and anatomical distribution of FLI during CTA expression. When animals received three rather than one conditioning trial, significant FLI was seen not only in the iNTS but also in the parabrachial nucleus (PBN), and the central nucleus of the amygdala (CNA), regions thought to be important in taste aversion learning.     

Effects of insular cortex lesions on conditioned taste aversion and latent inhibition in the rat

The present study tested the hypothesis that lesions of the insular cortex of the rat retard the acquisition of conditioned taste aversions (CTAs) because of an impairment in the detection of the novelty of taste stimuli. Demonstrating the expected latent inhibition effect, nonlesioned control subjects acquired CTAs more rapidly when the conditioned stimulus (0.15% sodium saccharin) was novel rather than familiar (achieved by pre-exposure to the to-be-conditioned taste cue). However, rats with insular cortex lesions acquired taste aversions at the same slow rate regardless of whether the saccharin was novel or familiar. The pattern of behavioural deficits obtained cannot be interpreted as disruptions of taste detection or stimulus intensity, but is consistent with the view that insular cortex lesions disrupt taste neophobia, a dysfunction that consequently retards CTA acquisition because of a latent inhibition-like effect.     

Conditioning method determines patterns of c-fos expression following novel taste-illness pairing

Conditioned taste aversions (CTAs) can be established by exposing rats to a novel taste CS through a bottle or through intra-oral (IO) infusion. Lesion studies suggest differences between the two methods in their engagement of brain circuits, as excitotoxic amygdala lesions have no effect on bottle-conditioned CTAs, but eliminate CTAs produced using IO infusion. Fos-like immunoreactivity (FLI) was used to compare patterns of brain activation after pairing CS taste and US drug using bottle and IO methods. Conditioning rats using the bottle method was associated with widespread elevations in FLI throughout the putative CTA circuit (basolateral and central nuclei of amygdala, insular cortex and nucleus of the solitary tract). In contrast, IO conditioning led to activation only in the central nucleus of amygdala. This supports the suggestion of differences in aversion processing as a function of conditioning method and may explain the greater reliance on amygdala of IO-conditioned CTAs due to engagement of a less distributed neural network.     

Conditioned taste aversion and amygdala lesions in the rat: a critical review

Studies using permanent lesions implicate the amygdala, a recipient of gustatory and viscerosensory information, in taste aversion learning. Reviewing this literature with respect to the location of the lesions and the quality of the behavioral methodology reveals little, if any, involvement of the medial amygdala or central nucleus in conditioned taste aversion. Although a disruption is found following damage to the basolateral region, the attenuated conditioned taste aversion appears to be a consequence of a lesion-induced impairment of neophobia rather than an association formation deficit. The key to understanding the functional significance of the basolateral amygdala in conditioned taste aversion reduces, we believe, to determining the role of this structure in gustatory neophobia.     

Brain-derived neurotrophic factor enhances conditioned taste aversion retention

Brain-derived neurotrophic factor (BDNF) has recently emerged as one of the most potent molecular mediators of not only central synaptic plasticity, but also behavioral interactions between an organism and its environment. Our previous studies on the insular cortex (IC), a region of the temporal cortex implicated in the acquisition and storage of conditioned taste aversion (CTA), have demonstrated that induction of long-term potentiation (LTP) in the projection from the basolateral nucleus of the amygdala (Bla) to the IC, previous to CTA training, enhances the retention of this task. Recently, we found that intracortical microinfusion of BDNF induces a lasting potentiation of synaptic efficacy in the Bla-IC projection of adult rats in vivo. In this work, we present experimental data showing that intracortical microinfusion of BDNF previous to CTA training enhances the retention of this task. These findings support the concept that BDNF may contribute to memory-related functions performed by a neocortical area, playing a critical role in long-term synaptic plasticity.     

In vivo effects of intracortical administration of NMDA and metabotropic glutamate receptors antagonists on neocortical long-term potentiation and conditioned taste aversion.

It has been proposed that long-term potentiation (LTP), a form of activity-dependent modification of synaptic efficacy, may be a synaptic mechanism for certain types of learning. Recent studies on the insular cortex (IC), a region of the temporal cortex implicated in the acquisition and storage of conditioned taste aversion (CTA), have demonstrated that tetanic stimulation of the basolateral nucleus of the amygdala (Bla) induce an N-methyl-D-aspartate (NMDA) dependent LTP in the IC of adult rats in vivo. Here we present experimental data showing that intracortical administration of the NMDA receptor competitive antagonists CPP (-3(-2 carboxipiperazin-4-yl)-propyl-1-phosphonic acid, 0.03 microg per hemisphere) and AP-5 (D(-)-2-amino-5-phosphonopentanoic, 2.5 microg per hemisphere) disrupt the acquisition of conditioned taste aversion, as well as IC-LTP induction in vivo. In contrast, administration of the metabotropic glutamate receptor antagonist MCPG ((RS)-alpha-methyl-4-carboxyphenylglycine, 2.5 microg per hemisphere) does not disrupt the acquisition of CTA nor IC-LTP induction. These findings are of particular interest since they provide support for the view that the neural mechanisms underlying NMDA-dependent neocortical LTP constitute a possible mechanism for the learning-related functions performed by the IC.     

c-Fos induction in response to saccharin after taste aversion learning depends on conditioning method

Increases in c-Fos-like immunoreactivity (FLI) in the intermediate nucleus of the solitary tract (iNTS) have been seen consistently as a correlate of the expression of a conditioned taste aversion (CTA) when conditioning occurs using taste delivery through intraoral (I/O) infusions. The present study examined whether a similar FLI response would occur when conditioning was accomplished by presenting the taste solution in a bottle. I/O and bottle methods generated aversions that were comparable, when judged by the behavioral response of solution rejection. However, elevations in FLI were seen only in animals conditioned with the I/O method. This finding adds to evidence that the neural pathways underlying CTA learning differ as a function of the type of conditioning method used.     

The NMDA receptor antagonist CPP impairs conditioned taste aversion and insular cortex long-term potentiation in vivo.

It has been proposed that long-term potentiation (LTP) a form of activity-dependent modification of synaptic efficacy, may be a synaptic mechanism for certain types of learning. Recent studies on the insular cortex (IC) a region of the temporal cortex implicated in the acquisition and storage of conditioned taste aversion (CTA), have demonstrated that tetanic stimulation of the basolateral nucleus of the amygdala (Bla) induce an N-methyl- -aspartate (NMDA) dependent LTP in the IC of adult rats in vivo. Here we present experimental data showing that intracortical administration of the NMDA receptor competitive antagonist CPP (-3(-2 carboxipiperazin-4-yl)-propyl-1-phosphonic acid) disrupts the acquisition of conditioned taste aversion, as well as, the IC-LTP induction in vivo. These findings are of particular interest since they provide support for the view that the neural mechanisms underlying NMDA dependent neocortical LTP, constitute a possible mechanism for the learning related functions performed by the IC.     

Differential participation of the NBM in the acquisition and retrieval of conditioned taste aversion and Morris water maze

Deficits in both learning and memory after lesions of the cholinergic basal forebrain, in particular the nucleus basalis magnocellularis (NBM), have been widely reported. However, the participation of the cholinergic system in either acquisition or retrieval of memory process is still unclear. In this study, we tested the possibility that excitotoxic lesions of the NBM affect either acquisition or retrieval of two tasks. In the first experiment, animals were trained for two conditioned taste aversion tasks using different flavors, saccharine and saline. The acquisition of the first task was before NBM lesions (to test retrieval) and the acquisition of the second task was after the lesions (to test acquisition). Accordingly, in the first part of the second experiment, animals were trained in the Morris water maze (MWM), lesioned and finally tested. In the final part of this experiment, another set of animals was lesioned, then trained in the MWM and finally tested. All animals were able to retrieve conditioned taste aversion (CTA) and MWM when learned before NBM lesions; however, lesions disrupted the acquisition of CTA and MWM. The results suggest that the NBM and cholinergic system may play an important role in acquisition but not during retrieval of aversive memories.     

Significance of cortical-amygdalar-hypothalamic connections in retention of conditioned taste aversion in rats

Rats were trained to avoid drinking 0.1 m NaCl solution by one to four pairings of this solution with an i.p. injection of 0.15 m LiCl during restricted periods of fluid access. The number of lickings of NaCl and nonpaired other solutions was measured during the preconditioning control period, after conditioning, and after lesions of various brain regions. The acquired learned aversion was abolished after bilateral lesions of the gustatory cortex, perirhinal cortex posterior to the gustatory cortex, or stria terminalis. Ablations of the frontal cortex had no effect. These results suggest that cortical-amygdalar-hypothalamic connections play a significant role in the retention of a conditioned taste aversion.     

Lipopolysaccharide dose dependently impairs rapid toxin (LiCl)-induced gustatory conditioning: a taste reactivity examination of the conditioned taste aversion

There is much debate on how immune activation affects cognitive processing. Research has shown that stimulation of the immune system can significantly impair, have no adverse effects, or enhance learning and memory processes in animals. The present experiment evaluated the effects of the bacterial endotoxin, lipopolysaccharide (LPS) on the acquisition of a rapidly acquired conditioned taste aversion using a toxin-containing food. Male Long Evans rats were fitted with intraoral cannulae and habituated to the taste reactivity procedure. Rats received two conditioning days, 72 h apart, in which they were injected systemically with LPS (200, 100, or 50 microg/kg) or NaCl (0.9% vehicle) and 90 min later placed in the taste reactivity test chamber. Rats were given 5 brief (1 min) intraoral infusions of either a LiCl-adulterated sucrose solution (0.15M LiCl+0.3M sucrose) or NaCl-sucrose solution (0.15M NaCl+0.3M sucrose) across a 1h period. On the test day (72 h after the last conditioning trial), rats were given a 2 min intraoral infusion of the respective taste in a drug-free state. Individual taste reactivity responses were recorded and analyzed. Results demonstrate that rats treated with LPS dose-dependently increased ingestive responding to the LiCl-sucrose flavor while at the same time showing reduced rejection response frequency on the two conditioning days. LPS treatment did not alter taste reactivity responding to the NaCl-sucrose solution. On the test day, the LPS groups again displayed a dose dependent increase in ingestive responses and a decrease in rejection responses to the LiCl-sucrose taste. The present results suggest that LPS-induced immune system activation, significantly impairs the rapid acquisition of a conditioned taste aversion.     

Critical role of insular cortex in taste but not odour aversion memory

Conditioned odour aversion (COA) and conditioned taste aversion (CTA) result from the association of a novel odour or a novel taste with delayed visceral illness. The insular cortex (IC) is crucial for CTA memory, and the present experiments sought to determine whether the IC is required for the formation and the retrieval of COA memory as it is for CTA. We first demonstrated that ingested odour is as effective as taste for single-trial aversion learning in rats conditioned in their home cage. COA, like CTA, tolerates long intervals between the ingested stimuli and the illness and is long-lasting. Transient inactivation of the IC during acquisition spared COA whereas it greatly impaired CTA. Similarly, blockade of protein synthesis in IC did not affect COA but prevented CTA consolidation. Moreover, IC inactivation before retrieval tests did not interfere with COA memory expression when performed either 2 days (recent memory) or 36 days after acquisition (remote memory). Similar IC inactivation impaired the retrieval of either recent or remote CTA memory. Altogether these findings indicate that the IC is not necessary for aversive odour memory whereas it is essential for acquisition, consolidation and retrieval of aversive taste memory. We propose that the chemosensory stimulations modulate IC recruitment during the formation and the retrieval of food aversive memory.