The alpha 3 subunit gene of the nicotinic acetylcholine receptor is a candidate gene for ethanol stimulation

Alcohol and nicotine are coabused, and preclinical and clinical data suggest that common genes may influence responses to both drugs. A gene in a region of mouse chromosome 9 that includes a cluster of three nicotinic acetylcholine receptor (nAChR) subunit genes influences the locomotor stimulant response to ethanol. The current studies first used congenic mice to confirm the influential gene on chromosome 9. Congenic F₂ mice were then used to more finely map the location. Gene expression of the three subunit genes was quantified in strains of mice that differ in response to ethanol. Finally, the locomotor response to ethanol was examined in mice heterozygous for a null mutation of the α3 nAChR subunit gene ( Chrna3 ). Congenic data indicate that a gene on chromosome 9, within a 46 cM region that contains the cluster of nAChR subunit genes, accounts for 41% of the genetic variation in the stimulant response to ethanol. Greater expression of Chrna3 was found in whole brain and dissected brain regions relevant to locomotor behavior in mice that were less sensitive to ethanol-induced stimulation compared to mice that were robustly stimulated; the other two nAChR subunit genes in the gene cluster (α5 and β4) were not differentially expressed. Locomotor stimulation was not expressed on the genetic background of Chrna3 heterozygous (+/−) and wild-type (+/+) mice; +/− mice were more sensitive than +/+ mice to the locomotor depressant effects of ethanol. Chrna3 is a candidate gene for the acute locomotor stimulant response to ethanol that deserves further examination.     

A Pair of Dopamine Neurons Target the D1-Like Dopamine Receptor DopR in the Central Complex to Promote Ethanol-Stimulated Locomotion in Drosophila

Dopamine is a mediator of the stimulant properties of drugs of abuse, including ethanol, in mammals and in the fruit fly Drosophila. The neural substrates for the stimulant actions of ethanol in flies are not known. We show that a subset of dopamine neurons and their targets, through the action of the D1-like dopamine receptor DopR, promote locomotor activation in response to acute ethanol exposure. A bilateral pair of dopaminergic neurons in the fly brain mediates the enhanced locomotor activity induced by ethanol exposure, and promotes locomotion when directly activated. These neurons project to the central complex ellipsoid body, a structure implicated in regulating motor behaviors. Ellipsoid body neurons are required for ethanol-induced locomotor activity and they express DopR. Elimination of DopR blunts the locomotor activating effects of ethanol, and this behavior can be restored by selective expression of DopR in the ellipsoid body. These data tie the activity of defined dopamine neurons to D1-like DopR-expressing neurons to form a neural circuit that governs acute responding to ethanol.     

NR2B‐deficient mice are more sensitive to the locomotor stimulant and depressant effects of ethanol

The NR2B subunit of N‐methyl d ‐aspartate glutamate receptors influences pharmacological properties and confers greater sensitivity to the modulatory effects of ethanol. This study examined behavioral responses to acute ethanol in a conditional knockout mouse model that allowed for a delayed genetic deletion of the NR2B subunit to avoid mouse lethality. Mice lacking the NR2B gene (knockout) were produced by mating NR2B[f/f] mice with CAMKIIa‐driven tTA transgenic mice and the tetO‐CRE transgenic mice. Adult male and female offspring representing each of the resultant genotypes (knockout, CAM, CRE and wildtype mice) were tested for open‐field locomotor activity following acute low‐ and high‐dose ethanol challenge as well as loss of righting reflex. Findings indicate that male and female mice lacking the NR2B subunit exhibited greater overall activity in comparison to other genotypes during the baseline locomotor activity test. NR2B knockout mice exhibited an exaggerated stimulant response to 1.5 g/kg (i.p.) and an exaggerated depressant response to 3.0 g/kg (i.p.) ethanol challenge. In addition, NR2B knockout mice slept longer following a high dose of ethanol (4.0 g/kg, i.p.). To evaluate pharmacokinetics, clearance rates of ethanol (1.5, 4.0 g/kg, i.p.) were measured and showed that female NR2B knockouts had a faster rate of metabolism only at the higher ethanol dose. Western blot analyses confirmed significant reduction in NR2B expression in the forebrain of knockout mice. Collectively, these data indicate that the NR2B subunit of the N‐methyl d ‐aspartate glutamate receptor is involved in regulating low‐dose stimulant effects of ethanol and the depressant/hypnotic effects of ethanol.     

Sensitivity to psychostimulants in mice bred for high and low stimulation to methamphetamine

Methamphetamine (MA) and cocaine induce behavioral effects primarily through modulation of dopamine neurotransmission. However, the genetic regulation of sensitivity to these two drugs may be similar or disparate. Using selective breeding, lines of mice were produced with extreme sensitivity (high MA activation; HMACT) and insensitivity (low MA activation; LMACT) to the locomotor stimulant effects of acute MA treatment. Studies were performed to determine whether there is pleiotropic genetic influence on sensitivity to the locomotor stimulant effect of MA and to other MA- and cocaine-related behaviors. The HMACT line exhibited more locomotor stimulation in response to several doses of MA and cocaine, compared to the LMACT line. Both lines exhibited locomotor sensitization to 2 mg/kg of MA and 10 mg/kg of cocaine; the magnitude of sensitization was similar in the two lines. However, the lines differed in the magnitude of sensitization to a 1 mg/kg dose of MA, a dose that did not produce a ceiling effect that may confound interpretation of studies using higher doses. The LMACT line consumed more MA and cocaine in a two-bottle choice drinking paradigm; the lines consumed similar amounts of saccharin and quinine, although the HMACT line exhibited slightly elevated preference for a low concentration of saccharin. These results suggest that some genes that influence sensitivity to the acute locomotor stimulant effect of MA have a pleiotropic influence on the magnitude of behavioral sensitization to MA and sensitivity to the stimulant effects of cocaine. Further, extreme sensitivity to MA may protect against MA and cocaine self-administration.     

Ethanol-induced activation and rapid development of tolerance may have some underlying genes in common

Ethanol exerts biphasic effects on behavior, stimulant at low doses and depressant at higher doses. In the present study we used two mouse genetic models to investigate the relationships among activating and depressant responses to alcohol. The first model was a panel of nine isogenic genotypes. FAST and SLOW mice, selectively bred for high and low ethanol-induced motor activation, respectively, were used as a second model. We used loss of righting reflex to assess initial sensitivity and acute functional tolerance to a hypnotic dose of ethanol (3 g/kg, 20% v/v). Blood ethanol concentration at the onset of loss of righting reflex was used as an estimate of initial sensitivity, while the difference between concentration values at the recovery and loss of righting represented an acute functional tolerance score. Mean initial sensitivity and acute functional tolerance values of the nine strains were correlated with a previously obtained measure of ethanol-induced locomotor activation. Activation correlated significantly with both initial sensitivity ( r _g = 0.80; P < 0.05) and acute functional tolerance ( r _g = 0.77; P < 0.05). Thus, inbred genotypes that were activated more by a low dose of ethanol were also more sensitive to and developed more acute tolerance to a high dose. FAST mice had initial sensitivity values similar to those of SLOW mice, but developed more pronounced tolerance, indicating that ethanol-induced activation and acute functional tolerance may be regulated by some common genetic mechanisms. In summary, these results supported a genetic association between ethanol-induced activation and rapid development of tolerance.     

Ethanol‐ and cocaine‐induced locomotion are genetically related to increases in accumbal dopamine

Neuroanatomical research suggests that interactions between dopamine and glutamate within the mesolimbic dopamine system are involved in both drug‐induced locomotor stimulation and addiction. Therefore, genetically determined differences in the locomotor responses to ethanol and cocaine may be related to differences in the effects of these drugs on this system. To test this, we measured drug‐induced changes in dopamine and glutamate within the nucleus accumbens (NAcc), a major target of mesolimbic dopamine neurons, using in vivo microdialysis in selectively bred FAST and SLOW mouse lines, which were bred for extreme sensitivity (FAST) and insensitivity (SLOW) to the locomotor stimulant effects of ethanol. These mice also show a genetically correlated difference in stimulant response to cocaine (FAST > SLOW). Single injections of ethanol (2 g/kg) or cocaine (40 mg/kg) resulted in larger increases in dopamine within the NAcc in FAST compared with SLOW mice. There was no effect of either drug on NAcc glutamate levels. These experiments indicate that response of the mesolimbic dopamine system is genetically correlated with sensitivity to ethanol‐ and cocaine‐induced locomotion. Because increased sensitivity to the stimulating effects of ethanol appears to be associated with greater risk for alcohol abuse, genetically determined differences in the mesolimbic dopamine response to ethanol may represent a critical underlying mechanism for increased genetic risk for alcoholism.     

Involvement of A2A receptors in anxiolytic,locomotor and motivational properties of ethanol in mice

We have shown previously that mice lacking the adenosine A_2A receptor (A_2AR) generated on a CD1 background self‐administer more ethanol and exhibit hyposensitivity to acute ethanol. We aimed to investigate if the increased propensity of A_2A^?/? mice to consume ethanol is associated with an altered sensitivity in the motivational properties of ethanol in the conditioned place preference (CPP) and conditioned taste aversion (CTA) paradigms and with an altered development of sensitization to the locomotor effects of ethanol. We also tested their sensitivity to the anxiolytic effects of ethanol. Our results show that A_2A^?/? mice produced on a CD1 background displayed a reduced ethanol‐induced CPP and an increased sensitivity to the anxiolytic and locomotor‐stimulant effects of ethanol, but they did not show alteration in ethanol‐induced CTA and locomotor sensitization. Ethanol‐induced CPP, ethanol consumption and the locomotor effects of ethanol were also tested in A_2A^?/? mice produced on a C57BL/6J background. Our results emphasized the importance of the genetic background because alteration in ethanol consumption and preference, ethanol‐induced CPP and locomotor‐stimulant effects were not found in knockout mice produced on the alcohol‐preferring C57BL/6J genetic background. Finally, the A_2AR agonist, 2‐p‐(2‐carboxyethyl)‐phenylethylamino‐5′‐N‐ethylcarboxamidoadenosine hydrochloride (CGS 21680), reduced ethanol consumption and preference in C57BL/6J mice. In conclusion, A_2AR deficiency in mice generated on a CD1 background leads to high ethanol consumption that is associated with an increased sensitivity to the locomotor‐stimulant/anxiolytic effects of ethanol and a decrease in ethanol‐induced CPP.     

JNK Pathway Activation Is Controlled by Tao/TAOK3 to Modulate Ethanol Sensitivity

Neuronal signal transduction by the JNK MAP kinase pathway is altered by a broad array of stimuli including exposure to the widely abused drug ethanol, but the behavioral relevance and the regulation of JNK signaling is unclear. Here we demonstrate that JNK signaling functions downstream of the Sterile20 kinase family gene tao/Taok3 to regulate the behavioral effects of acute ethanol exposure in both the fruit fly Drosophila and mice. In flies tao is required in neurons to promote sensitivity to the locomotor stimulant effects of acute ethanol exposure and to establish specific brain structures. Reduced expression of key JNK pathway genes substantially rescued the structural and behavioral phenotypes of tao mutants. Decreasing and increasing JNK pathway activity resulted in increased and decreased sensitivity to the locomotor stimulant properties of acute ethanol exposure, respectively. Further, JNK expression in a limited pattern of neurons that included brain regions implicated in ethanol responses was sufficient to restore normal behavior. Mice heterozygous for a disrupted allele of the homologous Taok3 gene (Taok3Gt) were resistant to the acute sedative effects of ethanol. JNK activity was constitutively increased in brains of Taok3Gt/+ mice, and acute induction of phospho-JNK in brain tissue by ethanol was occluded in Taok3Gt/+ mice. Finally, acute administration of a JNK inhibitor conferred resistance to the sedative effects of ethanol in wild-type but not Taok3Gt/+ mice. Taken together, these data support a role of a TAO/TAOK3-JNK neuronal signaling pathway in regulating sensitivity to acute ethanol exposure in flies and in mice.     

Genome-wide association for methamphetamine sensitivity in an advanced intercross mouse line

Sensitivity to the locomotor stimulant effects of methamphetamine (MA) is a heritable trait that utilizes neurocircuitry also associated with the rewarding effects of drugs. We used the power of a C57BL/6J × DBA/2J F(2) intercross (n = 676) and the precision of a C57BL/6J × DBA/2J F(8) advanced intercross line (Aap: B6, D2-G8; or F(8) AIL; n = 552) to identify and narrow quantitative trait loci (QTLs) associated with sensitivity to the locomotor stimulant effects of MA. We used the program QTLRel to simultaneously map QTL in the F(2) and F(8) AIL mice. We identified six genome-wide significant QTLs associated with locomotor activity at baseline and seven genome-wide significant QTLs associated with MA-induced locomotor activation. The average per cent decrease in QTL width between the F(2) and the integrated analysis was 65%. Additionally, these QTLs showed a distinct temporal specificity within each session that allowed us to further refine their locations, and identify one QTL with a 1.8-LOD support interval of 1.47 Mb. Next, we utilized publicly available bioinformatics resources to exploit strain-specific sequence data and strain- and region-specific expression data to identify candidate genes. These results illustrate the power of AILs in conjunction with sequence and gene expression data to investigate the genetic underpinnings of behavioral and other traits.     

Mouse ICSI with frozen-thawed sperm: the impact of sperm freezing procedure and sperm donor strain

Normal mouse offspring can be obtained from oocytes injected with frozen-thawed spermatozoa without cryoprotection, however, embryo development can be affected by sperm freezing procedure and sperm donor strain. In this study we observed that direct contact of mouse spermatozoa with liquid nitrogen did not affect their ability to activate injected oocytes but severely restricted subsequent in vitro embryo development to blastocyst stage. Tris-EDTA buffer and M2 were also shown to be better sperm freezing extenders than DPBS, allowing higher developmental potential. In addition, differences in embryo development obtained by intracytoplasmic sperm injection (ICSI) with frozen-thawed spermatozoa were observed between hybrid sperm donor strains. Frozen-thawed B6D2F1 spermatozoa provided higher embryo development than sperm cells from C57CBAF1.     

Effects of caffeine on social behavior, exploration and locomotor activity: interactions with ethanol

The effects of caffeine and its interaction with ethanol were examined in a test of social behavior and a holeboard test of exploration and locomotion. Male mice were injected i.p. with 15, 30 or 60 mg/kg caffeine alone or in combination with 2 g/kg ethanol. The animals were then put in pairs into a familiar arena, or examined individually in the holeboard. Only the highest dose of caffeine (60 mg/kg) had a significant effect on the time spent in social interaction and motor activity in the social behavior test: both measures were reduced. The duration and frequency of avoidance-irritability behavior was dose-dependently increased by caffeine. In the holeboard, caffeine caused a dose-dependent increase in locomotor activity. 30 mg/kg caffeine reversed the ethanol-induced reduction of time spent in social interaction, and 60 mg/kg caffeine antagonized the ethanol-induced increase in locomotor activity in both the social behavior and holeboard tests. Caffeine's effects on ethanol-induced behavioral changes are compared with those of other drugs.     

Influence of the biogenic amine tyramine on ethanol-induced behaviors in Drosophila

The biogenic amine tyramine has been implicated in drug-induced behavior. The Drosophila inactive mutant is characterized by reduced tyramine and octopamine levels and is defective in cocaine sensitization. To test whether there is an overlap in the use of the amine neurotransmitter system in ethanol- and cocaine-induced behaviors, mutant analyses were extended to the phenotypic characterization of inactive and other mutants effecting the tyramine and octopamine neurotransmitter system. The inactive mutant displays increased ethanol sensitivity and is impaired in the initial startle response upon ethanol application. Furthermore, this mutant fails to regulate its alcohol-induced hyperactivity properly. In contrast to the defects seen after cocaine application, inactive mutants develop normal ethanol tolerance and sensitize to the locomotor activating effect of ethanol. The tyramine-beta-hydroxylase mutant (TbetaH) with increased tyramine and depleted octopamine levels displays normal ethanol sensitivity, a startle repression, and hyperactivates more in response to ethanol. In addition, TbetaH mutants fail to develop a tolerance to the hyperactivating effect of ethanol. Ethanol-induced sensitization does not seem to be impaired in either mutant, suggesting that tyramine is not required for this process. The comparative analysis of the phenotypes associated with inactive and TbetaH mutants suggests that the fine tuning of ethanol-induced hyperactivity can be correlated with different tyramine levels. Defects in other aspects of ethanol-induced behaviors might be due to different molecules or mechanisms.     

The ethanol-induced open-field activity in rodents treated with isethionic acid, a central metabolite of taurine.

The effect of isethionic acid, a central metabolite of taurine, on ethanol-induced locomotor activity was investigated in rodents. Ten minutes following an (i.p.) simultaneous administration of ethanol (0.0, 1.5, 2.0, 2.5, 3.0, 3.5 g/kg) and isethionic acid (0.0, 22.5, 45.0, 90.0, 180.0 mg/kg), mice were placed in the open-field chambers and locomotor activity was measured during a ten-minute testing period. A significant interaction was found between isethionic acid and ethanol. Isethionic acid pre-treated mice (45.0, 90.0 and 180.0 mg/kg) showed a higher locomotor activity than the saline group at 2.5 and 3 g/kg of ethanol. In a second study, isethionic acid (45 mg/kg) and ethanol (1 g/kg) were simultaneously injected to rats. Ten minutes after the two treatments, rats were placed in the open-field chamber for a 30-minute period. The depressant effects that ethanol produced on rat locomotion were amplified by the same dose of isethionic acid as it affected ethanol-induced locomotion in mice (45 mg/kg). However, isethionic acid did not change the spontaneous locomotion at any of the doses tested in mice or rats. Since no differences in blood ethanol levels were detected in both mice and rats, the interaction between isethionic acid's action and ethanol-related locomotion does not seem to be due to different rates of absorption of ethanol or any other pharmacokinetic process related to ethanol levels. The current study displayed that isethionic acid, administered intraperitoneally, behaves in a similar way to its immediate precursor, taurine, by amplifying ethanol-induction of the locomotor activity.     

Sensitivity to the locomotor-stimulant effects of ethanol and allopregnanolone: a quantitative trait locus study of common genetic influence

Previous studies have suggested that common genetic mechanisms influence sensitivity to the locomotor-stimulant effects of ethanol and allopregnanolone. We conducted two quantitative trait locus (QTL) studies to identify chromosomal regions that harbor genes that influence locomotor response to ethanol (2 g/kg) and allopregnanolone (17 mg/kg) using F2 crosses between C57BL/6J and DBA/2J mice. Because our previous data from the BXD recombinant inbred strains had indicated that chromosome 2 contained QTL for sensitivity to the locomotor-stimulant effects of both ethanol and allopregnanolone, we also tested reciprocal chromosome 2 congenic strains for sensitivity to the locomotor-stimulant effects of both drugs. The F2 analysis for ethanol sensitivity identified significant QTL on chromosomes 1 and 2 and suggestive QTL on chromosomes 5 and 9. The analysis of the allopregnanolone F2 study identified suggestive QTL on chromosomes 3, 5 and 12. Suggestive evidence for a female-specific QTL on chromosome 2 was also found. The studies of congenic mouse strains indicated that both the congenic strains captured one or more QTL for sensitivity to the locomotor-stimulant effects of both ethanol (2 g/kg) and allopregnanolone (17 mg/kg). When Fisher's method was used to combine the P values for the RI, F2 and congenic studies of the chromosome 2 QTL, cumulative probability scores of 9.6 x 10(-15) for ethanol and 7.7 x 10(-7) for allopregnanolone were obtained. These results confirm the presence of QTL for ethanol and allopregnanolone sensitivity in a common region of chromosome 2 and suggest possible pleiotropic genetic influence on sensitivity to these drugs.     

The correlation between brain weight and behavior changes in response to ethanol administration in laboratory mice

The effects of ethanol injections on the F2 offspring of the cross between large-brain (LB) and small-brain (SB) mouse strains selected for high and low relative brain weights, respectively, have been studied. The parental strains have significantly differed in brain weight for many generations. The effects of ethanol (2.4 g/kg) have been compared in four subpopulations of mice that differ pairwise in brain weight. One pair of subpopulations has been isolated from the hybrid group and the other, from generation 22 of selection of the parental strains. The results of ANOVA have demonstrated that brain weight is related to the response to ethanol injections. The parameters of stereotyped behavior, which increased in after ethanol injections and reflected the decrease in exploratory activity) were different in mice with high and low relative brain weights. The pattern of behavioral changes after ethanol injections is the second (after increased learning ability) behavioral trait found to be correlated with brain weight.     

Role of dopamine D1-like receptors in methamphetamine locomotor responses of D2 receptor knockout mice

Behavioral sensitization to psychostimulants manifests as an increased locomotor response with repeated administration. Dopamine systems are accepted to play a fundamental role in sensitization, but the role of specific dopamine receptor subtypes has not been completely defined. This study used the combination of dopamine D2 receptor-deficient mice and a D1-like antagonist to examine dopamine D1 and D2 receptor involvement in acute and sensitized locomotor responses to methamphetamine. Absence of the dopamine D2 receptor resulted in attenuation of the acute stimulant effects of methamphetamine. Mutant and wild-type mice exhibited sensitization that lasted longer within the time period of the challenge test in the mutant animals. Pretreatment with the D1-like receptor antagonist SCH 23390 produced more potent reductions in the acute and sensitized locomotor responses to methamphetamine in D2 receptor-deficient mice than in wild-type mice; however, the expression of locomotor sensitization when challenged with methamphetamine alone was equivalently attenuated by previous treatment with SCH 23390. These data suggest that dopamine D2 receptors play a key role in the acute stimulant and sensitizing effects of methamphetamine and act in concert with D1-like receptors to influence the acquisition of methamphetamine-induced behavioral sensitization, traits that may influence continued methamphetamine use.     

Comparison of neurotensin levels, receptors and actions in LS/Ibg and SS/Ibg mice

Neurotensin (NT), injected centrally, markedly enhances sensitivity to ethanol-induced anesthesia in SS but not in LS mice (4). Since LS and SS mice were bred selectively for differential sensitivity to ethanol, these findings suggest that neurotensinergic neuronal processes mediate some of ethanol's actions and that LS and SS mice might differ genetically in neurotensinergic systems. Indeed, in biochemical studies it was shown that LS and SS mice differ in NT-like immunoreactivity in specific brain regions, i.e., hypothalamus, and in NT receptor densities (Bmax) in frontal cortex and striatum. In other experiments LS and SS mice differed in behavioral responses to centrally administered NT. Intracerebroventricular (ICV) administration of NT produced dose-dependent changes in motor activity, hypothermia, and analgesia in both LS and SS mice. SS mice appeared to be more sensitive than LS to NT-induced analgesia but not hypothermia. Neurotensin increased or decreased locomotor activity in both SS and LS mice following intraventral tegmental area or ICV administration, respectively. The results indicate that LS and SS mice, which were selectively bred for differences in ethanol sensitivity, differ genetically in NT concentrations, receptor densities in specific brain regions, and in some receptor-mediated behavioral responses to NT.     

Steroid-independent male sexual behavior in B6D2F2 male mice

It is well established that male sexual behavior (MSB) is regulated by gonadal steroids; however, individual differences in MSB, independent of gonadal steroids, are prevalent across a wide range of species, and further investigation is necessary to advance our understanding of steroid-independent MSB. Studies utilizing B6D2F1 hybrid male mice in which a significant proportion retain MSB after long-term orchidectomy, identified as steroid-independent-maters (SI-maters), have begun to unravel the genetic underpinnings of steroid-independent MSB. A recent study demonstrated that steroid-independent MSB is a heritable behavioral phenotype that is mainly passed down from B6D2F1 hybrid SI-maters when crossed with C57BL6J female mice. To begin to uncover whether the strain of the dam plays a role in the inheritance of steroid-independent MSB, B6D2F1 hybrid females were crossed with B6D2F1 hybrid males. While the present study confirms the finding that steroid-independent MSB is a heritable behavioral phenotype and that SI-mater sires are more likely to pass down some components of MSB than SI-non-maters to their offspring, it also reveals that the B6D2F2 male offspring that were identified as SI-maters that displayed the full repertoire of steroid-independent MSB had the same probability of being sired from either a B6D2F1 SI-mater or SI-non-mater. These results, in conjunction with previous findings, indicate that the specific chromosomal loci pattern that codes for steroid-independent MSB in the B6D2F2 male offspring may result regardless of whether the father was a SI-mater or SI-non-mater, and that the maternal strain may be an important factor in the inheritance of steroid-independent MSB.     

Thyroxine administration prevents matrilineal intergenerational consequences of in utero ethanol exposure in rats

The neurodevelopmental fetal alcohol spectrum disorder (FASD) is characterized by cognitive and behavioral deficits in the offspring. Conferring the deficits to the next generation would increase overall FASD disease burden and prevention of this transmission could be highly significant. Prior studies showed the reversal of these behavioral deficits by low dose thyroxine (T4) supplementation to the ethanol-consuming mothers. Here we aim to identify whether prenatal ethanol (PE) exposure impairs hippocampus-dependent learning and memory in the second-generation (F2) progeny, and whether T4 administration to the ethanol-consuming dam can prevent it. Sprague-Dawley (S) dams received control diets (ad libitum and nutritional control) or ethanol containing liquid diet with and without simultaneous T4 (0.3 mg/L diet) administration. Their offspring (SS F1) were mated with naive Brown Norway (B) males and females generating the SB F2 and BS F2 progeny. Hippocampus-dependent contextual fear memory and hippocampal expression of the thyroid hormone-regulated type 3 deiodinase, (Dio3) and neurogranin (Nrgn) were assessed. SS F1 PE-exposed females and their SB F2 progeny exhibited fear memory deficits. T4 administration to the mothers of F1 females reversed these deficits. Although SS F1 PE-exposed males also experienced fear memory deficit, this was neither transmitted to their BS F2 offspring nor reversed by prenatal T4 treatment. Hippocampal Dio3 and Nrgn expression showed similar pattern of changes. Grandmaternal ethanol consumption during pregnancy affects fear memory of the matrilineal second-generation progeny. Low dose T4 supplementation prevents this process likely via altering allele-specific and total expression of Dio3 in the hippocampus.