Age and experience dependent changes in Egr-1 expression during the ontogeny of the context preexposure facilitation effect (CPFE).

The context preexposure facilitation effect (CPFE) is a variant of contextual fear conditioning in which acquisition of the contextual representation and association of the retrieved contextual memory with an immediate foot-shock are separated by 24 h. During the CPFE, learning- related expression patterns of the early growth response-1 gene (Egr-1) vary based on training phase and brain sub-region in adult and adolescent rats (Asok, Schreiber, Jablonski, Rosen, & Stanton, 2013; Schreiber, Asok, Jablonski, Rosen, & Stanton, 2014; Chakraborty, Asok, Stanton, & Rosen, 2016). The current experiments extended our previous findings by examining Egr-1 expression in infant (PD17) and juvenile (PD24) rats during the CPFE using preexposure protocols involving single-exposure (SE) or multiple-exposure (ME) to context. Following a 5 min preexposure to the training context (i.e. the SE protocol), Egr-1 expression in the medial prefrontal cortex (mPFC), dorsal hippocampus (dHPC) and lateral nucleus of the amygdala (LA) was differentially increased in PD24 rats relative to PD17 rats. In contrast, increased Egr-1 expression following an immediate foot-shock (2s, 1.5 mA) did not differ between PD17 and PD24 rats, and was not learning-related. Interestingly, increasing the number of exposures to the training chamber on the preexposure day (i.e. ME protocol) altered training-day expression such that a learning-related increase in expression was observed in the mPFC in PD24 but not PD17 rats. Together, these results illustrate a clear maturation of Egr-1 expression that is both age- and experience-dependent. In addition, the data suggest that regional activity and plasticity within the mPFC on the preexposure but not the training day may contribute to the ontogenetic profile of the effect. Further studies are necessary to elucidate the causal role of sub-region-specific neuroplasticity in the ontogeny of the CPFE.

Antagonism of muscarinic acetylcholine receptors in medial prefrontal cortex disrupts the context preexposure facilitation effect.

Cholinergic function plays a role in a variant of context fear conditioning known as the context preexposure facilitation effect (CPFE; Robinson-Drummer, Dokovna, Heroux, & Stanton, 2016). In the CPFE, acquisition of a context representation, the context-shock association, and expression of context fear occur across successive phases, usually 24h apart. Systemic administration of scopolamine, a muscarinic acetylcholine receptor antagonist, prior to each phase (context preexposure, immediate-shock training, and testing) disrupts the CPFE in juvenile rats (Robinson-Drummer et al., 2016). Dorsal hippocampal (dHPC) cholinergic function contributes significantly to this effect, as local infusion of scopolamine into the dHPC prior to any individual phase of the CPFE produces a disruption identical to systemic administration (Robinson-Drummer et al., 2016). The current experiment extended these findings to another forebrain region implicated in the CPFE, the medial prefrontal cortex (mPFC). Adolescent rats received bilateral infusions of scopolamine (35µg/side) or PBS 10min before all three phases of the CPFE or only prior to a single phase. Intra-mPFC administration of scopolamine prior to all three phases significantly impaired fear conditioning suggesting that mPFC cholinergic function is necessary for successful CPFE performance. Analyses of the individual infusion days revealed a significant impairment of the CPFE when infusions occurred prior to preexposure or training (i.e. immediate footshock) but not prior to testing. In total, these findings suggests a role of mPFC cholinergic function in the acquisition and/or consolidation of a contextual representation and the context-shock association but not in retrieval or expression of fear memory. Implications for mPFC involvement in contextual fear conditioning and neurological dysfunction following neonatal alcohol exposure are discussed.

Variants of contextual fear conditioning induce differential patterns of Egr-1 activity within the young adult prefrontal cortex.

Contextual fear conditioning is a form of associative learning where animals must experience a context before they can associate it with an aversive stimulus. Single-trial contextual fear conditioning (sCFC) and the context preexposure facilitation effect (CPFE) are two variants of CFC where learning about the context is temporally contiguous (sCFC) with or separated (CPFE) from receiving a footshock in that context. Neural activity within CA1 of the dorsal hippocampus (CA1), amygdala (LA), and prefrontal cortex (PFC) may play a critical role when animals learn to associate a context with a footshock (i.e., training). Previous studies from our lab have found that early-growth-response gene 1 (Egr-1), an immediate early gene, exhibits unique patterns of activity within regions of the PFC following training in sCFC and the CPFE of juvenile rats. In the present study, we extended our studies by examining Egr-1 expression in young adult rats to determine (1) if our previous work reflected changes unique to development or extend into adulthood and (2) to contrast expression profiles between sCFC and the CPFE. Rats that learned context fear with sCFC showed increased Egr-1 in the anterior cingulate, orbitofrontal and infralimbic cortices relative to non-associative controls following training, but expression in prelimbic cortex did not differ between fear conditioned and non-associative controls. In contrast, rats trained in the CPFE also showed increased Egr-1 in all the prefrontal cortex regions, including prelimbic cortex. These findings replicate our previous findings in juveniles and suggest that Egr-1 in specific PFC subregions may be uniquely involved in learning context-fear in the CPFE compared to sCFC.

Egr-1 mRNA expression patterns in the prefrontal cortex, hippocampus, and amygdala during variants of contextual fear conditioning in adolescent rats.

We report activation of the immediate-early gene Egr-1 in the lateral amygdala (LA), hippocampus (CA1), and medial prefrontal cortex (mPFC) 30-min following the training phase in the context pre-exposure facilitation effect (CPFE) and standard context fear conditioning (180 s context exposure→shock). On day one of the CPFE paradigm, postnatal day (PD) 31 rats (±1) were pre-exposed to Context A (Pre) or Context B (Alt-Pre) for 5 min followed by five additional 1-min exposures. A day later, Pre and Alt-Pre rats received a 2-s, 1.5 mA footshock immediately upon placement in Context A. Animals included in in situ hybridization were then sacrificed 30 (±3) min later. On day three, the behaviorally-tested Pre rats showed significantly more fear-conditioned freezing in Context A than Alt-Pre rats. Standard context fear conditioning groups showed much greater freezing than the Pre group, as well as no shock and immediate-shock controls. Thirty minutes after immediate shock training, Pre rats showed increased Egr-1 mRNA in the prelimbic mPFC relative to Alt-Pre rats. Standard context conditioning selectively increased Egr-1 in CA1. In the LA and mPFC, Egr-1 increased to a similar extent in no shock, immediate shock, and standard context conditioning relative to homecage controls. The present study demonstrates that Egr-1 mRNA expression has a complex relationship to fear learning in different brain regions and variants of context conditioning.

Exercise and environment as an intervention for neonatal alcohol effects on hippocampal adult neurogenesis and learning.

Neonatal alcohol exposure impairs cognition and learning in adulthood and permanently damages the hippocampus. Wheel running (WR) improves hippocampus-associated learning and memory and increases the genesis and survival of newly generated neurons in the hippocampal dentate gyrus. WR significantly increases proliferation of newly generated dentate granule cells in alcohol-exposed (AE) and control rats on Postnatal Day (PD) 42 but only control rats show an increased number of surviving cells thirty days after WR (Helfer et al., 2009b). The present studies examined whether proliferation-promoting WR followed by survival-enhancing environmental complexity (EC) during adolescence could increase survival of new neurons in AE rats. On PD 4-9, pups were intubated with alcohol in a binge-like manner (5.25g/kg/day, AE), were sham-intubated (SI), or were reared normally (suckle control, SC). On PD 30 animals were assigned to WR (PD 30-42) followed by EC (PD 42-72; WR/EC) or were socially housed (SH/SH) for the duration of the experiment. All animals were injected with 200mg/kg bromodeoxyuridine (BrdU) on PD 41. In Experiment 1, survival of newly generated cells was significantly enhanced in the AE-WR/EC group in comparison with AE-SH/SH group. Experiment 2A examined trace eyeblink conditioning. In the SH/SH condition, AE impaired trace eyeblink conditioning relative to SI and SC controls. In the WR/EC condition, AE rats performed as well as controls. In Experiment 2B, the same intervention was examined using the context preexposure facilitation effect (CPFE); a hippocampus-dependent variant of contextual fear conditioning. Again, the WR/EC intervention reversed the deficit in conditioned fear to the context that was evident in the SH/SH condition. Post-weaning environmental manipulations promote cell survival and reverse learning deficits in rats that were exposed to alcohol during development. These manipulations may provide a basis for developing interventions that ameliorate learning impairments associated with human fetal alcohol spectrum disorders.

Neonatal alcohol impairs the context preexposure facilitation effect in juvenile rats: dose-response and post-training consolidation effects.

Alcohol exposure on postnatal days (PND) 4-9 in the rat adversely affects hippocampal anatomy and function and impairs performance on a variety of hippocampus-dependent tasks. Exposure during this developmental window reveals a linear relationship between alcohol dose and spatial learning impairment in the context preexposure facilitation effect (CPFE), a hippocampus-dependent variant of contextual fear conditioning. The purpose of the current report was to examine the effect of a range of alcohol doses administered during a narrower window, PND7-9, than previously reported (Experiment 1) and to begin to determine which memory processes involved in this task are impaired by developmental alcohol exposure (Experiment 2). In Experiment 1, rats pups received a single day binge alcohol dose of either 2.75, 4.00, 5.25 g/kg/day or were sham-intubated (SI) from PND7-9. Conditioned freezing during the test day was evident in all dosing groups, except for Group 5.25 g, indicating no graded dose-related behavioral deficits with alcohol exposure limited to PND7-9. In Experiment 2, rat pups were exposed to the highest effective dose from Experiment 1 (5.25 g/kg/day) or were sham intubated over PND7-9. During training, rats remained in the conditioning context for 5-min following immediate shock delivery. During this test of post-shock freezing, both SI and alcohol-exposed rats given prior exposure to the conditioning context showed comparable freezing levels. Since alcohol-exposed rats showed normal post-shock freezing, deficits by these rats on the test day likely reflect a failure to consolidate or retrieve a context-shock association, rather than a deficit in hippocampal conjunctive processes (consolidation, pattern completion) that occur prior to shock on the training day. These findings illustrate the value of the CPFE for characterizing the separable memory processes that are impaired by neonatal alcohol exposure in this task.

Determinants of novel object and location recognition during development.

In the novel object recognition (OR) paradigm, rats are placed in an arena where they encounter two sample objects during a familiarization phase. A few minutes later, they are returned to the same arena and are presented with a familiar object and a novel object. The object location recognition (OL) variant involves the same familiarization procedure but during testing one of the familiar objects is placed in a novel location. Normal adult rats are able to perform both the OR and OL tasks, as indicated by enhanced exploration of the novel vs. the familiar test item. Rats with hippocampal lesions perform the OR but not OL task indicating a role of spatial memory in OL. Recently, these tasks have been used to study the ontogeny of spatial memory but the literature has yielded conflicting results. The current experiments add to this literature by: (1) behaviorally characterizing these paradigms in postnatal day (PD) 21, 26 and 31-day-old rats; (2) examining the role of NMDA systems in OR vs. OL; and (3) investigating the effects of neonatal alcohol exposure on both tasks. Results indicate that normal-developing rats are able to perform OR and OL by PD21, with greater novelty exploration in the OR task at each age. Second, memory acquisition in the OL but not OR task requires NMDA receptor function in juvenile rats [corrected]. Lastly, neonatal alcohol exposure does not disrupt performance in either task. Implications for the ontogeny of incidental spatial learning and its disruption by developmental alcohol exposure are discussed.

Effects of exercise and environmental complexity on deficits in trace and contextual fear conditioning produced by neonatal alcohol exposure in rats.

In rodents, voluntary exercise and environmental complexity increases hippocampal neurogenesis and reverses spatial learning and long-term potentiation deficits in animals prenatally exposed to alcohol. The present experiment extended these findings to neonatal alcohol exposure and to delay, trace, and contextual fear conditioning. Rats were administered either 5.25 g/kg/day alcohol via gastric intubation or received sham-intubations (SI) between Postnatal Day (PD) 4 and 9 followed by either free access to a running wheel on PD 30-41 and housing in a complex environment on PD 42-72 (wheel-running plus environmental complexity; WREC) or conventional social housing (SHSH) from PD 30 to 72. Adult rats (PD 80 ± 5) received 5 trials/day of a 10-s flashing-light conditioned stimulus (CS) paired with .8 mA footshock either immediately (delay conditioning) or after a 10-s trace interval (trace conditioning) for 2 days. Neonatal alcohol exposure impaired context and trace conditioning, but not short-delay conditioning. The WREC intervention did not reverse these deficits, despite increasing context-related freezing in ethanol-exposed and SI animals.

Neonatal alcohol exposure and the hippocampus in developing male rats: effects on behaviorally induced CA1 c-Fos expression, CA1 pyramidal cell number, and contextual fear conditioning.

Rats exposed to a high binge-like dose of alcohol over postnatal days (PD) 4-9 show reductions in CA1 pyramidal cells and impairments on behavioral tasks that depend on the hippocampus. We first examined hippocampal c-Fos expression as a marker of neuronal activity in normally developing rats following different phases of the context preexposure facilitation effect (CPFE) paradigm (Experiment 1). During the CPFE, preexposure to the training context facilitates contextual conditioning to an immediate shock given on a subsequent occasion. We then examined the relationship between CPFE impairment, hippocampal cell loss, and c-Fos expression in rats exposed to alcohol over PD 4-9 (Experiment 2). Normally developing (Experiment 1), sham-intubated control (SI), and PD 4-9 alcohol-exposed (4.00 g and 5.25 g/kg/d; Experiment 2) juvenile male rats were trained on the CPFE. The CPFE occurs over three phases separated by 24 h. Starting on PD 31, rats were preexposed to Context A or Context B for 5 min. After 24 h, all rats received an immediate 1.5-mA foot shock in Context A. Finally, rats were tested for contextual conditioning in Context A on PD 33. Normally developing and SI rats preexposed to Context A showed enhanced contextual fear compared with those preexposed to Context B (Experiment 1) or alcohol-exposed rats preexposed to Context A (Experiment 2). Rats were sacrificed 2 h following different phases of the CPFE and processed for c-Fos immunohistochemistry (Experiments 1 and 2) and CA1 pyramidal cell quantification (Experiment 2). In Experiment 1, c-Fos positive (c-Fos+) cells in the dentate gyrus (DG) were consistently high among rats preexposed to Context A (Pre), Context B (No Pre), or sacrificed directly from their home cage (Home) and did not differ across CPFE phases. CA3 and CA1 c-Fos+ cells were highest during preexposure and decreased across training phases, with Group No Pre showing greater numbers of c-Fos+ cells during training than Group Pre and Controls. In Experiment 2, SI rats had greater numbers of CA1 c-Fos+ cells compared with alcohol-exposed rats, differing significantly from rats exposed to the high alcohol dose (5.25 g) over PD 4-9. Experiment 2 also revealed a linear decline in CA1 pyramidal cells across treatment groups, again with rats from the high-alcohol dose group showing significantly fewer CA1 pyramidal cells compared with SI. Our results reveal that context novelty may be a significant contributor to differential hippocampal c-Fos expression following different phases of the CPFE. In addition, lower levels of c-Fos+ cells in alcohol-exposed rats following preexposure may be related to general reductions in the number of CA1 pyramidal cells in these rats. The significant CPFE impairments in rats exposed to the lower alcohol dose (4.00 g), who show a 15% reduction in CA1 pyramidal cells compared with SI rats, highlight the sensitivity of the CPFE to hippocampal insult.

Neonatal alcohol exposure disrupts hippocampal neurogenesis and contextual fear conditioning in adult rats.

Developmental alcohol exposure can permanently alter brain structures and produce functional impairments in many aspects of behavior, including learning and memory. This study evaluates the effect of neonatal alcohol exposure on adult neurogenesis in the dentate gyrus of the hippocampus and the implications of such exposure for hippocampus-dependent contextual fear conditioning. Alcohol-exposed rats (AE) received 5.25g/kg/day of alcohol on postnatal days (PD) 4-9 (third trimester in humans), in a binge-like manner. Two control groups were included: sham-intubated (SI) and suckle-control (SC). Animals were housed in social cages (3/cage) after weaning. On PD80, animals were injected with 200mg/kg BrdU. Half of the animals were sacrificed 2h later. The remainder were sacrificed on PD114 to evaluate cell survival; separate AE, SI, and SC rats not injected with BrdU were tested for the context preexposure facilitation effect (CPFE; ~PD117). There was no difference in the number of BrdU+ cells in AE, SI and SC groups on PD80. On PD114, cell survival was significantly decreased in AE rats, demonstrating that developmental alcohol exposure damages new cells' ability to incorporate into the network and survive. Behaviorally tested SC and SI groups preexposed to the training context 24h prior to receiving a 1.5mA 2s footshock froze significantly more during the context test than their counterparts preexposed to an alternate context. AE rats failed to show the CPFE. The current study shows the detrimental, long-lasting effects of developmental alcohol exposure on hippocampal adult neurogenesis and contextual fear conditioning.

Factors governing single-trial contextual fear conditioning in the weanling rat.

Although contextual fear conditioning emerges later in development than explicit-cue fear conditioning, little is known about the stimulus parameters and biological substrates required at early ages. The authors adapted methods for investigating hippocampus function in adult rodents to identify determinants of contextual fear conditioning in developing rats. Experiment 1 examined the duration of exposure required by weanling rats at postnatal day (PND) 23 to demonstrate contextual fear conditioning. This experiment demonstrated that 30 s of context exposure is sufficient to support conditioning. Furthermore, preexposure enhanced conditioning to an immediate footshock, the context preexposure facilitation effect (CPFE), but had no effect on contextual conditioning to a delayed shock. Experiment 2 demonstrated that N-methyl-D-aspartate (NMDA) receptor inactivation during preexposure impairs contextual learning at PND 23. Thus, the conjuctive representations underlying the CPFE are NMDA-dependent as early as PND23 in the rat.

USAID program for the prevention and treatment of vaginal fistula.

The cornerstone of the US Agency for International Development (USAID) fistula program is to support and strengthen local capacity for fistula repair. The USAID program includes support to upgrade facilities, enhance local surgical repair capability, allocate equipment and supplies to operating rooms, implement quality improvement systems, and cover the women's transportation costs to and from the treatment facilities. The program also offers training in clinical and counseling skills; transferring skills South-to-South; and monitoring and evaluating the program's effectiveness. As new fistula cases continually increase the backlog of untreated cases, its efforts are also directed toward the prevention of fistula and the reintegration of treated women into their communities. Furthermore, the program challenges the culture of sexual violence against women that leads to traumatic gynecologic fistulas.

Effects of early hippocampal lesions on trace, delay, and long-delay eyeblink conditioning in developing rats.

The effects of bilateral hippocampal aspiration lesions on later acquisition of eyeblink conditioning were examined in developing Long-Evans rat pups. Lesions on postnatal day (PND) 10 were followed by evaluation of trace eyeblink conditioning (Experiment 1) and delay eyeblink conditioning (Experiment 2) on PND 25. Pairings of a tone conditioned stimulus (CS) and periocular shock unconditioned stimulus (US, 100 ms) were presented in one of three conditioning paradigms: trace (380 ms CS, 500 ms trace interval, 880 ms interstimulus interval [ISI]), standard delay (380 ms CS, 280 ms ISI), or long delay (980 ms CS, 880 ms ISI). The results of two experiments indicated that hippocampal lesions impaired trace eyeblink conditioning more than either type of delay conditioning. In light of our previous work on the ontogeny of trace, delay, and long-delay eyeblink conditioning (Ivkovich, Paczkowski, & Stanton, 2000) showing that trace and long-delay eyeblink conditioning had similar ontogenetic profiles, the current data suggest that during ontogeny hippocampal maturation may be more important for the short-term memory component than for the long-ISI component of trace eyeblink conditioning. The late development of conditioning over long ISIs may depend on a separate process such as protracted development of cerebellar cortex.

Perinatal exposure to polychlorinated biphenyls Aroclor 1016 or 1254 did not alter brain catecholamines nor delayed alternation performance in Long-Evans rats.

Several reports have indicated that polychlorinated biphenyls (PCB) altered development of biogenic amine systems in the brain, impaired behavioral performances, and disrupted maturation of the thyroid axis. The current study examines whether these developmental effects of PCB are correlated. Timed-pregnant Long-Evans rats were gavaged with the PCB mixture Aroclor 1016 (A-1016, 10 mg/kg) from gestation day (GD) 6 to parturition. Some pups continued to receive daily oral administration of PCB (10 mg/kg) until weaning at postnatal day (PD) 21. Another group of pregnant rats was given Aroclor 1254 (A-1254, 8 mg/kg) daily from GD 6 to weaning. At various age intervals, rats were sacrificed and six brain regions (prefrontal cortex, striatum, hippocampus, diencephalon, cerebellum, midbrain + brain stem) were removed and analyzed for dopamine (DA) and norepinephrine (NE) levels by high-performance liquid chromatography. In addition, transmitter turnover rates were determined after an acute treatment of alpha-methyl-p-tyrosine. Serum samples were collected and analyzed for triiodothyronine (T(3)) and thyroxine (T(4)) by radioimmunoassay. Behaviorally, rats were evaluated for spatial learning and memory by means of T-maze delayed alternation and Morris maze tasks on PD 23 and PD 70, respectively. A-1016 treatment produced only small and transient reductions in body weight gain, and generally did not alter the thyroid status of the developing rats. It did not cause any significant changes in DA or NE level, or turnover rate in any of the brain regions examined, nor did it affect behavioral measures of cognitive development. In contrast, perinatal exposure to A-1254 led to marked deficits of growth, and sharply reduced serum T(4), although T(3) remained largely unaffected. Accompanying this hormonal imbalance, brain NE contents in the A-1254-exposed pups were reduced, although brain DA was not significantly affected; no demonstrable neurobehavioral deficits were seen in the T-maze or Morris maze tests. These results indicated that development of central noradrenergic neurons was compromised by perinatal exposure to A-1254 but not A-1016, and both PCB mixtures failed to alter behavioral performances.

Ontogenetic differences in the effects of unpaired stimulus preexposure on eyeblink conditioning in the rat.

Learned irrelevance (LIr) is a Pavlovian conditioning phenomenon in which random or unpaired preexposure to a conditional stimulus (CS) and to an unconditional stimulus (US) retards subsequent paired conditioning involving these stimuli. A previous developmental study of eyeblink conditioning in the rat suggested that LIr is not present on postnatal Day 20. Stanton, Fox, and Carter (1998) showed that unpaired preexposure to a CS and a US on postnatal Day 17 failed to retard (and, in fact, facilitated) subsequent paired conditioning involving these stimuli on postnatal Day 20. The present experiments were designed to further characterize the ontogeny of this phenomenon. In Experiment 1, LIr was observed when rat pups were tested for eyeblink conditioning as described in Stanton et al. (1998), except that preexposure occurred on postnatal Day 27, and acquisition testing occurred on postnatal Day 30. In Experiment 2, preexposure and acquisition both occurred on postnatal Day 30, and four types of preexposure were compared: chamber only, CS alone, US alone, or unpaired presentation of CS and US. Unpaired preexposure impaired acquisition relative to that of the remaining three groups, which did not differ. Experiment 3, showed that under the conditions of Experiment 2, LIr failed to appear on postnatal Day 20, but was observed on postnatal Days 25 and 30. These findings suggest that learning that events are unrelated emerges between postnatal Days 20 and 25 in the rat. Possible behavioral and neural mechanisms underlying this effect are discussed.

Multiple memory systems, development and conditioning.

A century of behavioral and neurobiological research suggests that Pavlovian conditioning involves three component memory systems: sensorimotor, affective and cognitive. In classical eyeblink conditioning, there is evidence that these three memory systems involve, respectively, the cerebellum, amygdala and hippocampus. This article reviews developmental research on eyeblink conditioning in rodents that is beginning to characterize ontogenetic dissociations and interactions among these memory systems. This research shows that the functional development of the affective system (conditioned fear response) precedes that of the sensorimotor system (conditioned eyeblink reflex). Modulation of these two systems by cognitive processes also seems to emerge at different points in ontogeny. Implications for cognitive development and research on multiple memory systems are discussed.

Ontogeny of delay versus trace eyeblink conditioning in the rat.

The ontogeny of delay versus trace eyeblink conditioning was examined in 19-, 23-, and 30-day-old rat pups. Pairings of a tone conditioned stimulus (CS) and periocular shock unconditioned stimulus (US; 100-ms) were presented in one of three conditioning paradigms: standard delay [380-ms CS, 280-ms interstimulus interval (ISI)], trace (380-ms CS, 500-ms trace interval), or long-delay (980-ms CS, 880-ms ISI). The results of two experiments indicated that standard delay conditioning emerged between 19 and 23 days of age whereas trace and long-delay eyeblink conditioning emerged more slowly from postnatal Days 19 to 30. Because the acquisition profile for long-delay paralleled that of trace and not standard delay, it appears that the relative deficits in the emergence of trace eyeblink conditioning during development reflect difficulty in forming associations over long ISIs rather than the short-term memory demands of the trace conditioning paradigm.

Ontogeny of eyeblink conditioning using a visual conditional stimulus.

The developmental emergence of associative learning in rodents is determined by interactions among sensory, motor, and associative systems that are engaged in a particular experimental preparation (Carter & Stanton, 1996; Hunt & Campbell, 1997; Rudy, 1992). In fear conditioning, chemosensory, auditory, and visual cues emerge successively as effective conditional stimuli (CS) during postnatal ontogeny. In the present study, we begin to examine the generality of this principle of sensory system development for eyeblink conditioning, a form of associative learning that develops substantially later than conditioned fear (Carter & Stanton, 1996). We asked whether the developmental emergence of eyeblink conditioning to a visual CS occurs at an age that is the same or different from conditioning to an auditory CS. In Experiment 1, rat pups were trained on postnatal Day 17 or 24 with experimental parameters (and design) that were identical to our previous studies of eyeblink conditioning except that presentation of a light rather than a tone served as the CS. The outcome was also identical: no eyeblink conditioning on Day 17 and strong conditioning on Day 24. In Experiment 2, conditioning to tone versus light was directly compared by means of a discrimination learning design on postnatal Days 19, 21, 23, and 31. There was no evidence for differential development of auditory versus visual eyeblink conditioning. The difference between this outcome and previous ones involving conditioned fear (Hunt & Campbell, 1997; Rudy, 1992) suggests that principles concerning sensory maturation and learning may be different for early- versus late-developing associative systems.

Development of cholinergic neurons in rat brain regions: dose-dependent effects of propylthiouracil-induced hypothyroidism.

The effects of hypothyroidism on development of cholinergic system in brain regions (prefrontal cortex and hippocampus) were evaluated by measuring choline acetyltransferase (ChAT) activity and hemicholinium-3 binding to the high-affinity choline transporter. Various degrees of thyroid deficiency were produced by perinatal exposure to propylthiouracil (PTU) in drinking water ranging from 5 ppm (mg/l) to 25 ppm beginning at gestational day 18 until postnatal day 21. ChAT, a marker for cholinergic nerve terminals, was reduced by PTU in a dose-dependent manner. Concomitant with the enzyme deficits, hemicholinium-3 binding was elevated, suggesting an increase in neuronal impulse activity. Although similar changes were seen in both brain regions examined, the magnitude and duration of these changes were more definitive in the prefrontal cortex. Nonetheless, these neurochemical alterations appeared to be recoverable when the rats returned to a euthyroid state, and no further changes were observed as the animals reached adulthood. In comparison, data reported in a succeeding article indicate that deficits in cognitive function were first seen in weanling hypothyroid rats, but that the behavioral impairments lasted well into adulthood when thyroid status and cholinergic parameters in the brain appeared to have recovered to normal. These results suggest that alterations of cholinergic system caused by perinatal hypothyroidism are associated with neurobehavioral deficits at weaning, and these developmental deviations may cause permanent impairment of cognitive function despite recovery from the hormonal imbalance at adult ages.