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Keywords:

  • methamphetamine;
  • HPA axis, corticosterone;
  • arginine vasopressin, glucocorticoid receptor;
  • sex difference

Abstract

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Results
  5. Discussion
  6. Acknowledgements and conflict of interest disclosure
  7. References
Thumbnail image of graphical abstract

Dysregulation of hypothalamic–pituitary–adrenal (HPA) axis activation is associated with changes in addiction-related behaviors. In this study, we tested whether sex differences in the acute effects of methamphetamine (MA) exposure involve differential activation of the HPA axis. Male and female mice were injected with MA (1 mg/kg) or saline for comparison of plasma corticosterone and analysis of the immediate early gene c-Fos in brain. There was a prolonged elevation in corticosterone levels in female compared to male mice. C-Fos was elevated in both sexes following MA in HPA axis-associated regions, including the hypothalamic paraventricular nucleus (PVN), central amygdala, cingulate, and CA3 hippocampal region. MA increased the number of c-Fos and c-Fos/glucocorticoid receptor (GR) dual-labeled cells to a greater extent in males than females in the cingulate and CA3 regions. MA also increased the number of c-fos/vasopressin dual-labeled cells in the PVN as well as the number and percentage of c-Fos/GR dual-labeled cells in the PVN and central amygdala, although no sex differences in dual labeling were found in these regions. Thus, sex differences in MA-induced plasma corticosterone levels and activation of distinct brain regions and proteins involved in HPA axis regulation may contribute to sex differences in acute effects of MA on the brain.

Methamphetamine induces a prolonged plasma corticosterone response in females compared to males. This may be mediated by increased neural activation, involving a greater activation of glucocorticoid receptor-positive cells, in males in the CA3 and cingulate brain regions, which are involved in negative feedback functions. These findings indicate a sex difference in the neural regulation of methamphetamine-induced plasma corticosterone release.

Abbreviations used
AVP

arginine vasopressin

BNST

bed nucleus of the stria terminalis

CRH

corticotropic-releasing hormone

GR

glucocorticoid receptor

PBS

phosphate-buffered saline

PVN

paraventricular nucleus of the hypothalamus

PVT

paraventricular thalamic nucleus

In the United States, methamphetamine (MA) was reported to be abused by approximately 439 000 people in 2011, 133 000 of which were new users (SAMHSA 2012). MA abuse results in devastating consequences for those who use the drug as well as for the society as a whole. MA is estimated to cost the United States approximately $23.4 billion annually (Nicosia et al. 2009). There are sex differences in the use and subsequent effects of MA. Women begin using MA at an earlier age than men (Dluzen and Liu 2008) and have a shorter duration from first use to regular use (Brecht et al. 2004). Furthermore, women are more dependent on and committed to MA (SAMHSA 2010), whereas men tend to use other drugs when MA is unavailable to them (Brecht et al. 2004). Rodent models also indicate that females show greater MA seeking and reinstatement of MA use following a period of withdrawal (Roth and Carroll 2004; Reichel et al. 2012).

Chronic hypothalamic–pituitary–adrenal (HPA) axis activation can lead to lasting alterations in HPA axis function (Herman et al. 1995; Gómez et al. 1996). Dysregulation of HPA axis function is strongly linked to changes in emotional (e.g., depression, anxiety) as well as addiction-related behaviors including MA seeking (Shoener et al. 2006; Fernández-Guasti et al. 2012; Nawata et al. 2012). Both emotional and addictive behaviors have been reported to be sexually dimorphic in humans and rodents (Palanza 2001; Brecht et al. 2004; Leach et al. 2008; Reichel et al. 2012) although the contribution of MA-induced activation of the HPA axis is unknown. In rodents, MA exposure in the developing brain has been shown to acutely and chronically activate the HPA axis, which results in increased plasma levels of adrenocorticotropic hormone and corticosterone (Williams et al. 2000; Acevedo et al. 2008). In the developing brain, the effects of MA exposure on HPA axis activation are more pronounced in female than male mice (Acevedo et al. 2008). In contrast to the developing brain, little is known about potential sex differences in HPA axis activation following MA exposure in adulthood which may contribute to sex differences in MA effects on the brain and behavior.

Activation of the HPA axis involves a complex brain circuitry. This circuitry includes areas of the cortex, amygdala, hippocampus, and hypothalamus, which project to the paraventricular nucleus of the hypothalamus (PVN), the central HPA axis regulatory nucleus in the brain. Within the PVN, several cell phenotypes are critical for the regulation of the HPA axis, including arginine vasopressin (AVP), corticotropic-releasing hormone (CRH), and glucocorticoid receptor (GR)-expressing cells. AVP-expressing cells within the PVN are divided into two functionally distinct subpopulations, magnocellular and parvocellular neurons. These neurons increase transcription of AVP following HPA axis activation (Angulo et al. 1991). Magnocellular AVP-expressing cells project to the posterior pituitary and are involved in the regulation of water and electrolyte homeostasis (Skorecki et al. 1992) as well as cardiovascular function (Robertson 1977). AVP synthesized from the parvocellular population acts in conjunction with CRH to stimulate secretion of adrenocorticotropic hormone from the pituitary gland, and subsequently, glucocorticoids (e.g., corticosterone) from the adrenal gland (Plotsky 1987). Following activation of the HPA axis, glucocorticoids activate GRs located throughout the brain and pituitary. Within the brain, activation of GRs in the cortex, hippocampus, amygdala, hypothalamus, and other regions are involved in terminating the HPA axis through negative feedback mechanisms (Herman et al. 2012). GR activation also contributes to stress-related memory formation (de Kloet et al. 2005) and has been linked to addiction-related behaviors (Wang et al. 2008; Ambroggi et al. 2009). In contrast, glucocorticoids primarily act via mineralocorticoid receptors in the brain to regulate glucocorticoid levels during basal conditions (Reul and de Kloet 1985; Dallman et al. 1987).

Activation of the HPA axis and corticosterone release have also been linked to addiction-related behaviors. Chronic elevations in plasma corticosterone levels lead to alterations in HPA axis-associated genes and proteins within the PVN, amygdala, and hippocampus (Kovács and Mezey 1987; Reul et al. 1987; Herman et al. 1989; Sawchenko et al. 1993). Disruptions in these genes, particularly CRH, have been shown to affect MA addiction-related behaviors including MA seeking following withdrawal (Nawata et al. 2012).

In conditions of stress in adulthood, female rodents show greater increases in plasma corticosterone levels and different activation patterns in HPA axis-associated brain regions compared to males (Handa et al. 1994a; Figueiredo et al. 2002; Zuloaga et al. 2011). As little is known about potential sex differences in HPA axis activation following MA exposure in adulthood; in this study, we investigated and compared MA-induced plasma corticosterone responses in male and female mice. MA also increases body temperature for a prolonged period of time (Kuo et al. 2003) which coincides with activation of the HPA axis, although the specific relationship between the two is not well understood. Therefore, we also examined core body temperature responses to MA at the same sampling time points for plasma corticosterone to examine the temporal relationship between these measures. In addition, we explored immediate early gene activation in regions associated with the HPA axis to assess potential MA and sex differences in activation of brain nuclei. Finally, we analyzed the colocalization patterns of the immediate early gene c-Fos with AVP and GR to identify the specific cell phenotypes activated by MA. These phenotypes were chosen based on previous studies that indicate MA can modify their expression (Zuloaga et al., 2013, Lowy 1990). We hypothesized that females would show elevated corticosterone responses to MA and an altered activation of HPA axis-associated brain regions compared to males.

Materials and methods

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Results
  5. Discussion
  6. Acknowledgements and conflict of interest disclosure
  7. References

Animals

C57BL6J mice were purchased from JAX Laboratories (Bar Harbor, ME, USA) and maintained on a 12/12 light/dark cycle with lights on at 6:00 am. Mice were singly housed for 1 week prior to experiments and were between 60–90 days old at the time of testing. Rodent chow (PicoLab Rodent Diet 20, #5053; PMI Nutrition International, St. Louis, MO, USA) and water were available ad libitum.

Blood collection, body temperature measurement, and corticosterone assay

For corticosterone analysis, male and female mice were injected with (d)-MA hydrochloride {[obtained from the Research Triangle Institute (Research Triangle Park, NC) through the National Institute on Drug Abuse drug supply program]} dissolved in saline (1 mg/kg, ip.) or saline alone. Trunk blood was collected at 30, 70, or 120 min after injection (n = 6 per treatment/sex). Trunk blood was also collected from singly housed male (n = 6) and female mice (n = 6) who received no injection to obtain baseline measures. All blood collection occurred between 8:00–11:00 am. Following no injection, or at the specified time points after the saline or MA injection, mice were removed from their cage and body temperatures were measured using a Microtherma 2 rectal thermometer (Thermoworks, Lindon, UT, USA). Immediately following temperature measurement, mice were killed by cervical dislocation followed by rapid decapitation and collection of trunk blood into EDTA-treated tubes. Blood was centrifuged at 5500 g for 10 min and the supernatant was transferred to a new tube and stored at −80°C until assay. Plasma samples were analyzed using commercial I125 corticosterone radioimmunoassays according to the manufacturer's instructions (MP Biochemicals, LLC, Orangeburg, NY, USA). Intra- and interassay coefficients of variation were 4.4% and 7.3%, respectively.

c-Fos Immunohistochemistry

For c-Fos immunohistochemical analysis, male and female mice were injected with (d)-MA hydrochloride dissolved in saline (1 mg/kg, i.p.), saline alone, or not injected between 10:00–11:00 am (n = 5 per treatment/sex, except n = 4 for the no injection male group). Two hours later, mice were intracardially perfused with 20-mL phosphate-buffered saline (PBS) followed by 40 mL 4% paraformaldehyde. Brains were removed, stored in 4% paraformaldehyde overnight, and then transferred to 30% sucrose. The 2-hour time point was chosen based on previous studies demonstrating extensive induction of c-Fos and corticosterone in the mouse brain following administration of the same dose of MA (Zhu et al. 2010; Giardino et al. 2011). Fixed brains were coronally sectioned at 40 μm into three series using a cryostat (Microm HM505E, MICROM international GmbH, Walldorf, Germany) and processed for immunohistochemical detection of c-Fos. Sections were rinsed in PBS, incubated in 1% hydrogen peroxide and 0.3% Triton-X in PBS (PBS-TX) for 10 min, again rinsed in PBS, then incubated in 10% normal goat serum in PBS-TX for 1 h. After rinsing in PBS, sections were incubated in primary antisera (c-Fos rabbit polyclonal: 1 : 5000, Santa Cruz Biotechnology, sc52, Billerica, MA, USA) in 4% normal goat serum and PBS-TX overnight at 21°C. Sections were then rinsed in PBS and incubated for 1 h in biotinylated goat-anti-rabbit antibody in PBS-TX (1 : 500; Vector Laboratories, Burlingame, CA, USA) followed by rinses in PBS and a 1-h incubation in avidin–biotin peroxidase complex (ABC Elite kit; Vector Laboratories). Following rinses in tris-buffered saline, sections were developed for visualization of c-Fos-positive cells in a hydrogen peroxide/diaminobenzidine/tris-buffered saline solution for 10 min, after which sections were rinsed in PBS and immediately mounted on slides. The following day, sections were dehydrated in ethanol, defatted in xylene, and coverslipped with Permount (Sigma Chemical Co., St. Louis MO, USA).

Dual-Label Immunohistochemistry

To determine the colocalization of c-Fos and AVP or GR immunoreactivity in the mouse brain, we performed dual-label immunohistochemistry. For c-Fos/AVP or c-Fos/GR double labeling, free-floating sections were rinsed with PBS three times, then blocked with 4% donkey serum in PBS-TX for 90 min. Sections were then incubated in anti-c-Fos (1 : 250, goat, Santa Cruz Biotechnology, sc52-G) overnight. Sections were then incubated in 1 : 200 donkey anti-goat Dylight 594 (Abcam, Cambridge, MA, USA) for 3 h at 21°C. Sections were then rinsed in PBS four times (20 min each rinse) after which the same protocol was repeated using GR (1 : 250, rabbit, M-20, Santa Cruz Biotechnology) or AVP (1 : 250, rabbit, gift from Dr Paul Plotsky) as primary antibody and 1 : 200 donkey anti-rabbit Alexa 488 (Life Technologies, Grand Island, NY, USA) as the secondary antibody. Sections were slide mounted and coverslipped with anti-fade reagent to preserve fluorescent signal (Vectashield with 4′,6-diamidino-2-phenylindole (DAPI), Vector Laboratories), light protected, and stored at 4°C. Brain sections incubated without primary antibody were used as negative controls to test for c-Fos, vasopressin, and GR-ir and showed no labeling.

Microscopy

Quantification of c-Fos positive cells was performed using an Olympus IX81 microscope (Olympus, Center Valley, PA, USA) equipped with Slidebook software (Intelligent Imaging Innovations, Inc., Denver, CO, USA). Brain regions were identified using the mouse brain atlas of Franklin and Paxinos (2007). Images of the PVN (Bregma ≈ −0.82 and −0.94 mm), paraventricular thalamic nucleus (PVT; Bregma ≈ −0.82 and −0.94 mm), bed nucleus of the stria terminalis (BNST; Bregma ≈ 0.02 and −0.10 mm), central nucleus of the amygdala (CEA; Bregma ≈ −1.34 and −1.46 mm), cingulate cortex area 2 (Bregma ≈ 1.10 and 0.98 mm), dentate gyrus, CA1, and CA3 hippocampal regions (Bregma ≈ −0.82 and −0.94 mm) were captured bilaterally within two sections using a 10× objective. c-Fos-immunoreactive cells (identified by black nuclear label) were quantified bilaterally within fixed area frames; PVN (box, 275 × 450 μm), PVT (box, 790 × 410 μm each), BNST (box, 335 × 620 μm), CEA (circle, 575 μm diameter), cingulate cortex (box, 425 × 400 μm), dentate gyrus (box, 850 × 420 μm), CA1 (box, 850 × 420 μm), and CA3 (box, 850 × 420 μm).

For quantification of c-Fos/GR and c-Fos/AVP dual labeling, confocal images of the PVN (c-Fos/GR and c-Fos/AVP), CEA, CA3, and cingulate (c-Fos/GR) were captured bilaterally at 20× in two sections using the same Olympus IX81 confocal microscope. GR, AVP, c-Fos, and dual-labeled cells were quantified in the CEA (GR; circle, 275 μm diameter), CA3 (GR; box, 430 × 300 μm), cingulate (GR; box, 350 × 270 μm), and separately within lateral magnocellular [(GR and AVP) circle, 135 μm diameter] and medial parvocellular regions of the PVN [(GR and AVP) circle, 135 μm diameter]. Cells were considered colocalized when they expressed both red (c-Fos) and green (AVP or GR) fluorescence. For AVP/c-fos coexpressing cells AVP was found within the extra-nuclear area, whereas c-Fos was localized to the nucleus, resulting in a green cell body that surrounds a red nucleus. GR labeling was found in the nucleus, so c-Fos/GR coexpressing cells were visualized as yellow/orange resulting from an overlap of the red and green secondary labeling. Within brain regions, c-Fos, GR, and AVP cells were counted as were cells coexpressing c-Fos/GR or c-Fos/AVP. These counts were then used to determine both the total number and percentage of c-Fos-positive cells that expressed GR or AVP. For all cell quantifications, cells were counted in both hemispheres of a given region in each section and summed. Cell counts for two sections were averaged and are presented as cells per section.

Statistical analysis

All data are reported as means ± SEM. Data were analyzed using GraphPad Prism v.4 and SPSS v.16.0 software (Chicago, IL, USA). For the immunohistochemical analyses, data are presented as the number and percentage of c-Fos-positive cells expressing GR or AVP. For corticosterone and body temperature analysis, three-way anova was utilized with treatment (MA, saline), sex (male, female), and time as between subject factors (0, 30, 70, 120 min). Immunohistochemical data (c-Fos, c-Fos/AVP, and c-Fos/GR) were analyzed using two-way anova with sex and treatment as factors as well as separated by sex and analyzed as fold change compared to saline injection. Correlations between corticosterone and body temperature were performed using Spearman's Rank Correlation Coefficient. Bonferroni corrected post hoc comparisons were performed when statistically appropriate. Student's t-tests were utilized for comparison of fold increases in c-Fos and c-Fos/GR dual-labeled cells in MA compared to saline-treated mice.

Results

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Results
  5. Discussion
  6. Acknowledgements and conflict of interest disclosure
  7. References

Corticosterone

A three-way anova revealed a significant main effect of treatment (F(1,77) = 70.48, < 0.001). Mice treated with MA showed increased plasma corticosterone levels compared to saline-treated mice (Fig. 1a). There was also a significant main effect of sex (F(1,77) = 11.741, p = 0.001), with greater plasma corticosterone levels in females than males. Finally, there was a main effect of time (F(3,77) = 57.96, p < 0.001) and a significant treatment × sex × time interaction (F(3,77) = 3.874, p = 0.012). This interaction reflected significantly elevated plasma levels of corticosterone in MA-treated females at 70 and 120 min.

image

Figure 1. Plasma corticosterone and body temperature responses to methamphetamine exposure. (a) Compared to saline, methamphetamine (MA) injection resulted in an elevated plasma corticosterone response in male and female mice. Female mice exhibited a prolonged rise in plasma corticosterone levels compared to males following MA exposure, with elevated levels at 70 and 120 min. (b) Injection with MA or saline resulted in a significant elevation in body temperature at 30 min. At 70 min and 120 min body temperature continued to rise in MA-treated mice, whereas in saline-treated mice it declined. Correlation between the plasma corticosterone response and body temperature revealed a positive correlation in saline-injected (c) and a negative correlation in MA-injected (d) mice. *indicates a significant treatment × sex × time interaction resulting from a prolonged rise in plasma corticosterone levels in female compared to male MA-treated mice. #indicates a significant increase in body temperature in MA- compared to saline-treated mice.

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Body temperature

A three-way anova revealed a significant main effect of treatment (F(1,77) = 20.62, p < 0.001). Mice treated with MA showed an elevated temperature compared to saline-treated mice (Fig. 1b). There was also a significant main effect of time (F(3,77) = 45.62, p < 0.001), indicating elevations in temperature following treatment with saline or MA. There was a significant treatment × sex interaction (F(3,77) = 4.951, p = 0.029) and a treatment × time interaction (F(3,77) = 13.70, p < 0.001). Saline and MA treatments had sex-dependent effects on body temperature, particularly at the 70-min time point, in which MA induces a more robust elevation in body temperature in males than females compared to same-sex saline mice. The effects of saline and MA differed in terms of duration of body temperature elevation. Specifically, MA induced a prolonged elevation in body temperature compared to saline treatment.

Next, we assessed correlations of plasma corticosterone and body temperature at 30, 70, and 120 min following MA or saline injection. These correlations did not differ by sex. Therefore, the data of female and male mice were combined for correlational analyses. For saline-treated mice, there was a significant positive correlation (ρ(35)=0.445; p = 0.007) (Fig. 1c) between body temperature and plasma corticosterone levels, but in MA-treated mice there was a negative correlation between body temperature and plasma corticosterone levels (ρ(35)= −0.564; p < 0.001; Fig. 1d). This negative correlation appeared driven by the maintained elevation in body temperature occurring in opposition to declining plasma corticosterone levels at later time points.

c-Fos quantification

There was a significant main effect of treatment on the number of c-Fos-immunopositive cells in the PVN ((F(2,20)= 25.12, p < 0.001; Fig. 2a), CEA ((F(2,20) = 40.01, p < 0.001; Fig. 2d), BNST ((F(2,20) = 31.78, p < 0.001; Fig. 2e), PVT ((F(2,20) = 48.68, p < 0.001; Fig. 2b), cingulate ((F(2,20) = 31.74, p < 0.001; Fig. 3a), CA1 ((F(2,20) = 21.95, p < 0.01; Fig. 2c), and CA3 ((F(2,20) = 9.52, p < 0.01; Fig. 3c). There was an overall increase in the number of c-Fos-immunoreactive neurons in MA-treated compared to saline-treated (p < 0.01) or no injection control groups (p < 0.01). Furthermore, several regions including the PVN, PVT, and BNST also showed increased numbers of c-Fos-positive cells following saline injection compared to the no injection groups (p < 0.01). This is likely because of an injection stress-induced activation of these brain regions. Only the dentate gyrus (DG) showed no significant effect of treatment (Fig. 2f). For most brain regions, there was no significant difference between males and females in the number of c-Fos-ir neurons, either at baseline or in response to saline or MA injection. However, the fold increase in c-Fos-positive cells in MA-treated compared to saline-treated mice was greater in males within the cingulate cortex (t(8) = 4.70; p = 0.0015; Fig. 3b) and CA3 region of the hippocampus (t(8) = 3.34; p = 0.0103; Fig. 3d).

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Figure 2. Effect of methamphetamine on c-Fos expression in hypothalamic–pituitary–adrenal (HPA) axis-associated brain regions. (a) Administration of 1 mg/kg MA caused a significant increase in the expression of c-Fos in the (a) PVN, (b) PVT, (c) CA1, (d) CEA, and (e) bed nucleus of the stria terminalis (BNST) compared to no injection and saline controls. (f) MA did not alter c-Fos expression in the dentate gyrus. Saline-injected mice also showed a significant increase in c-Fos expression in the (a) PVN and (b) PVT compared to mice that were not injected. (f) Representative images of methamphetamine effects on regional c-Fos expression. Rectangles and spheres approximate shapes used for quantification. Images were captured using a 10× objective. *indicates p < 0.001 compared to no injection and saline groups; #indicates p < 0.01 compared to no injection group. MA, methamphetamine; PVN, paraventricular hypothalamic nucleus; CEA, central amygdala; PVT, paraventricular thalamic nucleus; BNST, bed nucleus of the stria terminalis; DG, dentate gyrus.

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Figure 3. Methamphetamine causes sex-specific increases in c-Fos in the cingulate cortex and CA3 hippocampal region. (a) Treatment with 1 mg/kg MA resulted in a significant increase in the expression of c-Fos in the cingulate cortex compared to no injection and saline controls. Female mice showed fewer c-Fos-positive cells following MA when compared to males. Saline-injected mice also showed a significant increase in the number of c-Fos expressing cells in the cingulate compared to mice that were not injected. (b) Compared to saline-treated mice of the same sex, males showed a greater induction of c-Fos-positive cells in the cingulate compared to females. (c) MA resulted in a significant increase in c-Fos expression in CA3 compared to no injection and saline controls. Female mice showed fewer c-Fos-positive cells following MA when compared to males. (d) Compared to saline-treated mice of the same sex, males showed a greater induction of c-Fos-positive cells compared to females in CA3. (e) Representative images of methamphetamine effects on c-Fos expression in the CA3 hippocampal region and cingulate cortex in male and female mice. Images were captured using a 10× objective. Rectangles approximate shapes used for quantification. *indicates p < 0.001 compared to no injection and saline groups; #indicates p < 0.01 compared to no injection group. *indicates p < 0.01 compared to no injection and saline groups; #indicates p < 0.01 compared to no injection group; ^indicates significant difference compared to MA-treated males; **indicates p ≤ 0.01 compared to males.

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c-Fos dual labeling

c-Fos/GR dual labeling was quantified in the two regions which indicated sex differences in total c-Fos (CA3 and cingulate). In both, the CA3 and cingulate regions, there was a main effect of treatment. There was a significant increase in the number of c-Fos/GR dual-labeled cells in MA-treated mice {CA3: (F(1,16) = 23.2, p < 0.001); Cingulate: (F(1,16) = 76.6, p < 0.001; Fig. 4a,d)}. Both regions also revealed a significant interaction between treatment and sex {CA3: (F(1,16) = 4.74, p = 0.045); Cingulate: (F(1,16) = 5.27, p = 0.035)} indicating a greater MA induction in c-Fos/GR colocalized cells in males compared to females. Post hoc analysis also revealed a significantly greater number of c-Fos/GR coexpressing cells in male compared to female MA-treated mice in the cingulate (p < 0.05). The fold increase in c-Fos/GR-positive cells from saline-treated to MA-treated mice was greater in males within the cingulate cortex (t(8) = 3.96; p = 0.004; Fig. 4b) and CA3 (t(8)=3.83; p = 0.005; Fig. 4e). No significant differences were found for the percentage of c-Fos cells coexpressing GR in either region (Fig. 4c,f). There were also no significant sex or treatment differences in the number of GR-positive cells or in the percentage of GR-positive cells that expressed c-Fos.

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Figure 4. Colocalization of c-Fos with glucocorticoid receptor (GR) in the cingulate cortex and CA3 hippocampal region following treatment with methamphetamine (MA) or saline. (a) MA significantly increased the number of c-Fos/GR dual-labeled cells in the cingulate compared to saline controls. Female mice showed fewer c-Fos/GR-positive cells following MA when compared to males. (b) Compared to saline-treated mice of the same sex, males showed a greater induction of c-Fos/GR-positive cells in the cingulate compared to females. (c) No differences were found in the percentage of c-Fos-positive cells that expressed GR in the cingulate. (d) MA resulted in a significant increase in c-Fos/GR dual-labeled cells in CA3 compared to saline controls. (e) Compared to saline-treated mice of the same sex, males showed a greater induction of CA3 c-Fos/GR-positive cells compared to females. (f) No differences were found in the percentage of c-Fos-positive cells that expressed GR in CA3. *indicates p < 0.01 compared to saline groups; ^indicates significant difference compared to MA-treated males; **indicates p < 0.01 compared to males.

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C-Fos dual labeling was also quantified within two regions central to HPA axis regulation, the PVN and CEA. The total number of c-Fos/GR dual-labeled cells was greater in both the parvocellular (F(1,16) = 81.04, p < 0.001; Fig. 5a) and magnocellular (F(1,16) = 56.15, p < 0.001; Fig. 5e) PVN regions as well as the CEA (F(1,16) = 74.8, p < 0.001; Fig. 5i) following MA compared to saline treatment. Furthermore, the percentage of c-Fos-positive cells expressing GR was also significantly increased by MA treatment in the parvocellular PVN (F(1,16) = 21.41, p < 0.001; Fig. 5b), magnocellular PVN (F(1,16) = 17.98, p < 0.001; Fig. 5f), and CEA (F(1,16) = 59.2, p < 0.001; Fig. 5j) as compared to saline treatment. There were no sex differences in c-Fos/GR colocalized cells in the PVN or CEA. The total number of c-Fos/AVP dual-labeled cells was greater in both the parvocellular (F(1,16) = 9.23, p < 0.01; Fig. 5c) and magnocellular (F(1,16) = 15.06, p < 0.01; Fig. 5g) PVN regions, although the percentage of c-Fos-positive cells expressing AVP did not significantly differ by treatment in either PVN area (Fig. 5d,h). There were no significant sex differences in c-Fos/AVP colocalized cells. There were also no significant sex or treatment differences in the number of GR- or AVP-positive cells or in the percentage of GR-positive cells that expressed c-Fos. See Fig. 6 for representative images of c-Fos/GR and c-Fos/AVP dual-labeled cells.

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Figure 5. Colocalization of c-Fos with arginine vasopressin (AVP) and glucocorticoid receptor (GR) in the PVN and CEA following treatment with MA or saline. (a) The number c-Fos/GR dual-labeled cells and (b) the percentage of c-Fos cells that coexpress GR were increased following MA treatment in the PVNp. (c) The number c-Fos/AVP dual-labeled cells was increased following MA while (d) the percentage of c-Fos cells that coexpress AVP did not differ by treatment in the PVNp. (e) The number c-Fos/GR dual-labeled cells and (f) the percentage of c-Fos cells that coexpress GR was increased following MA treatment in the PVNm. (g) The number c-Fos/AVP dual-labeled cells was increased following MA, whereas (h) the percentage of c-Fos cells that coexpress AVP did not differ by treatment in the PVNm. (i) The number c-Fos/GR dual-labeled cells and (j) the percentage of c-Fos cells that coexpress GR were increased following MA treatment in the CEA. *indicates p < 0.01, **indicates p < 0.001 compared to saline treatment. PVNp, parvocellular paraventricular hypothalamic nucleus; PVNm, magnocellular paraventricular hypothalamic nucleus; CEA, central amygdala.

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Figure 6. Representative images of c-Fos dual labeling with arginine vasopressin (AVP) or glucocorticoid receptor (GR) in the methamphetamine-treated mouse PVN. (a) c-fos/AVP dual labeling in the PVN at low magnification. Circles indicate representative areas used to quantify labeling in the magnocellular (upper right circle) and parvocellular (lower left circle) PVN. Square indicates the region further magnified in (b–d). c-Fos/GR dual labeling in the PVN (e), CEA (i), CA3 (m), and cingulate (q) at low magnification. Circles indicate representative areas utilized to quantify labeled cells in the magnocellular (upper right circle) and parvocellular (lower left circle) PVN. Square indicates the region of PVN, CEA, CA3, and cingulate further magnified in (f–h, j–l, n–p, and r–t), respectively. Purple arrows indicate cells that express AVP or GR only. Gray arrows indicate cells that express c-Fos only. Blue arrows indicate cells that express both AVP/c-Fos or GR/c-Fos.

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Discussion

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Results
  5. Discussion
  6. Acknowledgements and conflict of interest disclosure
  7. References

The results of this study indicate that MA causes sex-dependent activation of the HPA axis with females showing a more prolonged elevation in plasma corticosterone compared to males. In females, the plasma corticosterone response following MA exposure peaked at 70 min post injection and subsequently decreased, but remained elevated above baseline levels at 120 min. In males, MA induced an earlier peak plasma corticosterone response at 30 min with a faster decline by 70 min and levels similar to baseline at 120 min. This sex difference in HPA axis activation with greater plasma corticosterone responses in females than males is similar to that observed in adult rodents following novelty and restraint stress (Handa et al. 1994a, 2009; Zuloaga et al. 2011), as well as administration of pharmacological agents such as the SSRI citalopram (Goel et al. 2011). Furthermore, the pattern of sex difference in HPA axis activation seen in the current study is similar to that seen following chronic exposure to MA from post-natal days 11–20 in the developing mouse brain (Acevedo et al. 2008). Saline injection alone induced a smaller rise in plasma corticosterone levels which peaked in male and female mice at 30 min and declined thereafter. This finding is consistent with previous studies indicating that stress from injection itself increases plasma corticosterone levels in mice (Goel et al. 2011).

The higher plasma corticosterone responses to MA in females than males may be regulated by sex differences in endogenous androgen and estrogen levels and differential activation of androgen and estrogen receptors. Gonadectomy of male rodents increases, while treatment with testosterone or dihydrotestosterone decreases, the plasma corticosterone response to stress (Handa et al. 1994b). Opposite effects are found in females, in which gonadectomy decreases and treatment with estradiol increases, plasma corticosterone responses to stress (Handa et al. 2009). These sex-dependent effects are mediated via estrogen receptor alpha stimulatory, and androgen receptor and estrogen receptor beta suppressive effects on the HPA axis (Zuloaga et al. 2008; Handa et al. 2009; Weiser and Handa 2009). Whether MA acts through similar hormone and receptor pathways to regulate sex differences in corticosterone release has yet to be determined.

Unlike plasma corticosterone responses, no sex differences were found in the MA-induced rise in body temperature. This temperature rise replicated a previous study which reported comparable rises in body temperature in male and female mice following MA administration (Kuo et al. 2003). Saline injection itself also resulted in a rise in body temperature, likely caused by stress-induced hyperthermia (McGivern et al. 2009). These data are comparable to previous reports, in which stress induced by injections resulted in temporally similar elevations and declines in plasma corticosterone levels and body temperature (Veening et al. 2004). However, MA and saline injections resulted in opposite correlations between plasma corticosterone levels and body temperature. There was a negative correlation between plasma corticosterone levels and body temperature in MA-treated mice. This negative correlation seemed driven by the maintained elevation in body temperature occurring in opposition to declining plasma corticosterone levels at later time points. This finding therefore indicates that there are distinct mechanisms that regulate HPA axis activation and hyperthermia following MA exposure. Activation of both independent and overlapping brain circuitry contribute to regulation of these functions following stress (Veening et al. 2004; Busnardo et al. 2010). Specifically, the PVN appears to be critical to the regulation of both functions (Veening et al. 2004; Busnardo et al. 2010). Other regions including the dorsomedial hypothalamus, anterodorsal preoptic area, and periaqueductal gray may be more pertinent to control of body temperature changes that parallel activation of the HPA axis (Veening et al. 2004). However, specific effects of MA on neural control of body temperature and the HPA axis are less understood.

Sex differences in MA activation of the HPA axis may be related to increased MA seeking in females. The corticosterone response to MA and MA seeking are both greater in female than male rodents (Roth and Carroll 2004; Reichel et al. 2012). One possibility is that the prolonged release of corticosterone in females may lead to greater alterations in HPA axis-associated genes and proteins. In turn this may contribute to alterations in MA seeking and reinstatement following withdrawal since disruptions in these genes, particularly CRH, have been linked to MA addiction-related behaviors (Nawata et al. 2012). Further studies are needed to test this hypothesis and determine the molecular pathways involved in the bidirectional relationship between effects of MA on HPA axis-associated proteins and effects of alterations in the levels and/or activational states of these proteins on MA addiction.

This increased HPA axis reactivity to MA in female mice is paralleled by a decrease in c-Fos-positive cells in two distinct brain regions, the CA3 region of the hippocampus and cingulate cortex. A sex difference in c-Fos induction in the CA3 area with greater activation in males than females has been reported following restraint stress (Figueiredo et al. 2002) and following administration of the SSRI citalopram (Goel et al. 2011), which also activates the HPA axis (Jensen et al. 1999). In both studies, decreased plasma corticosterone responses paralleled increased c-Fos expression in the CA3 region. The CA3 hippocampal region plays a key role in negative feedback functions of the HPA axis (Silverman et al. 1981). Therefore, activation of cells in this region may contribute to reducing the HPA axis response following its initial activation. In support of this notion, electrical stimulation of the dorsal CA3 region reduces plasma corticosterone levels (Dunn and Orr 1984). MA also increased c-Fos immunoreactivity within the CA1 hippocampal region, although not in a sex-dependent fashion, and failed to alter expression in the dentate gyrus. This suggests that sex differences in hippocampus-associated negative feedback of the HPA axis may be conferred primarily through the hippocampal CA3 area.

MA exposure also resulted in a sexually dimorphic c-Fos response in the cingulate cortex, again with males showing a greater activation. Sex differences in c-Fos activation in this brain region have previously been reported following stress, in which males also show greater responses than females in a similar division of the cingulate cortex (Figueiredo et al. 2002). Like the CA3 area, the cingulate is part of the neural circuitry that regulates inhibition of HPA axis function via multisynaptic projections to the PVN (Silverman et al. 1981; Diorio et al.1993). Reduced GR/Fos dual labeling in both the CA3 and cingulate cortex further suggest a mechanism for decreased negative feedback since activation of GR in hippocampal and cortical regions is critical to this function (Furay et al. 2008). Sex differences in neural activation within the cingulate and CA3 may also indicate differential effects of MA on memory functions and stress-related behaviors that are linked these brain regions (Handelmann and Olton 1981; Hunsaker et al. 2009; Wang et al. 2010). CA3 has also been linked to withdrawal-like behaviors and may potentially contribute to sex differences in this behavior in rodents (Isaacson and Lanthorn 1981). However, the specific role of CA3 in MA withdrawal is currently unknown.

MA administration also induced c-Fos expression in several other HPA axis-associated brain regions including the PVN, CEA, BNST, and PVT. In the PVN and CEA, two regions central to HPA axis activation, there were large inductions of c-Fos-positive cells following MA which parallel elevations in plasma corticosterone levels. However, no sex differences were found in MA-induced c-Fos-positive cells within these brain regions, although the plasma corticosterone response was greater in females. This finding is similar to reports in rats and mice that also demonstrate that c-Fos expression in the PVN and CEA does not differ although plasma corticosterone levels are elevated in females (Figueiredo et al. 2002; Campbell et al. 2003; Goel et al. 2011). Specifically, sex differences in corticosterone response (females > males) do not correlate with elevated c-Fos expression in females in these studies. It is possible that c-Fos expression in the PVN and CEA may be sexually dimorphic at different time points, either earlier or later, and involve either more or less activation of the same number of specific cells, or recruitment of more or less cells. However, in the current study we assessed only one specific time point at which previous studies have indicated extensive brain c-Fos levels following activation of the HPA axis (Giardino et al. 2011; Zuloaga et al. 2011). Further studies are needed to assess potential sex differences in the temporal dynamics of c-Fos following MA exposure.

The PVT and BNST also showed large inductions of c-Fos following MA injection with no sex differences. The PVT is proposed to be a key component of the brain circuitry that regulates habituation and facilitation of HPA axis function following stress (Bhatnagar and Dallman 1998; Bhatnagar et al. 2002), although this role has been debated (Fernandes et al. 2002). Given the high level of brain activation in the PVT following MA, it is possible that the PVT regulates similar HPA axis functions in the context of MA. The PVT projects to both the PVN and CEA and may therefore mediate HPA axis functions via these connections (Sawchenko and Swanson 1983; Turner and Herkenham 1991). The BNST has been demonstrated to play an inhibitory role on the HPA axis via GABAergic projections to the PVN (Choi et al. 2007). Therefore, activation of cells within this region may confer negative feedback regulation of the HPA axis. Robust neural activation of these brain regions (PVN, CEA, BNST, PVT) may also mediate MA effects on stress-related behaviors associated with these brain regions, particularly those associated with anxiety and depression (Salomé et al. 2004; Ventura-Silva et al. 2013). Long-term consequences of MA on these brain regions and their effects on stress and addiction-associated behaviors remain to be determined.

In an attempt to identify the phenotype of cells activated within the PVN and CEA, we performed dual-label immunohistochemistry to determine colocalization of c-Fos-positive cells with two proteins associated with the HPA axis, AVP and glucocorticoid receptor. Our results indicate that acute MA exposure results in a selective activation of neurons expressing GR with a lesser activation of AVP containing cells. This extensive activation of GR-positive cells in the PVN and CEA following MA exposure is greater than that following injection stress both in terms of total number of c-Fos/GR-positive cells and in the percentage of c-Fos-positive cells coexpressing GR. The percentage of c-Fos/GR-positive cells following acute MA injection reported here is also greater than that reported in rats subjected to restraint (Zavala et al. 2011), indicating that MA activation of GR-positive cells may be even greater than that observed following stress. However, species differences in HPA axis function may also contribute to this difference. Activation of GR has been linked to addiction-related behaviors (Wang et al. 2008; Ambroggi et al. 2009) although little is known about the specific involvement of the PVN and CEA in this process. In this study, we demonstrate that GR-positive cells in the PVN and CEA are specifically responsive to MA and may therefore be targets for MA-induced behavioral alterations. In the current study, we focused on GR rather than MR given their more prominent role in regulating HPA axis negative feedback in response to high levels of corticosterone (Reul and de Kloet 1985). However, the balance of GR/MR activation and expression in the brain is critical to HPA axis regulation (De Kloet and Derijk, 2004). Therefore, it is likely that MR also contributes to the corticosterone response following MA.

A smaller number and percentage of c-Fos-positive cells also expressed AVP, suggesting a lesser role for this neuropeptide in acute MA-induced activation of the HPA axis. It is possible that AVP may play a greater role in chronic activation of the HPA axis following repeated MA exposure. AVP has been reported to be involved in the adaptation of the HPA axis to chronic stress (Volpi et al. 2004). CRH-, rather than AVP-, expressing neurons in the PVN may be more specifically involved in MA activation of the HPA axis. Unfortunately, labeling for CRH in the mouse PVN is difficult to detect using available primary antisera, therefore studies utilizing alternative methods for visualizing CRH (eg. CRH-GFP models) are warranted to explore activation of these cells following MA exposure. C-Fos colocalization patterns with both AVP and GR did not differ between males and females suggesting that differential activation of these phenotypes in the PVN and CEA are not involved in sex differences in corticosterone response to MA. However, it remains possible that these cells may be activated for varying lengths of time in males compared to females thus resulting in a differential corticosterone response. Alternatively, a greater activation of CRH-positive cells in the PVN female mice may also contribute to their elevated corticosterone response. There is also some overlap in the distribution of GR and AVP-ir cells within the PVN (Uht et al., 1988; de Souza and Franci, 2010) so it is possible that activation of cells expressing both phenotypes may differ in response to MA. Further studies are warranted to test these possibilities.

In summary, the results of this study indicate that adult female mice show a prolonged corticosterone response to MA compared to males. Furthermore, we identified several HPA axis-associated regions that are activated following MA exposure. Two of these regions that have been associated with negative feedback regulation of the HPA axis (CA3 and cingulate) display a greater number of c-Fos and c-Fos/GR-positive cells after MA exposure in males than females. This suggests that these regions may contribute to sex differences in MA-induced corticosterone release. Finally, we report that MA injection results in extensive activation of GR-positive cells in the PVN and CEA with a lesser activation of AVP-positive cells in the PVN. Therefore, these cell phenotypes, particularly GR, appear to regulate effects of MA on the HPA axis and may therefore be potential targets for modulating MA effects on HPA axis function. These findings provide insight into anatomical regions and cell phenotypes involved in MA activation of the HPA axis and may also contribute to increased understanding of mechanisms involved in sex differences in MA use and the consequences of MA use.

Acknowledgements and conflict of interest disclosure

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Results
  5. Discussion
  6. Acknowledgements and conflict of interest disclosure
  7. References

Funding was provided by NIDA T32DA007262, an Oregon Health and Science University Tartar Award, and the Oregon Health and Science University development account of Dr Raber.

All experiments were conducted in compliance with the ARRIVE guidelines. The authors have no conflict of interest to declare.

References

  1. Top of page
  2. Abstract
  3. Materials and methods
  4. Results
  5. Discussion
  6. Acknowledgements and conflict of interest disclosure
  7. References
  • Acevedo S. F., Pfankuch T., van Meer P. and Raber J. (2008) Role of histamine in short- and long-term effects of methamphetamine on the developing mouse brain. J. Neurochem. 107, 976986.
  • Ambroggi F., Turiault M., Milet A. et al. (2009) Stress and addiction: glucocorticoid receptor in dopaminoceptive neurons facilitates cocaine seeking. Nat. Neurosci. 12, 247249.
  • Angulo J. A., Ledoux M. and McEwen B. S. (1991) Genomic effects of cold and isolation stress on magnocellular vasopressin mRNA-containing cells in the hypothalamus of the rat. J. Neurochem. 56, 20332038.
  • Bhatnagar S. and Dallman M. F. (1998) Neuroanatomical basis for facilitation of hypothalamic-pituitary-adrenal responses to a novel stressor after chronic stress. Neuroscience 84, 10251039.
  • Bhatnagar S., Huber R., Nowak N. and Trotter P. (2002) Lesions of the posterior paraventricular thalamus block habituation of hypothalamic-pituitary-adrenal responses to repeated restraint. J. Neuroendocrinol. 14, 403410.
  • Brecht M. L., O'Brien A., von Mayrhauser C. and Anglin M. D. (2004) Methamphetamine use behaviors and gender differences. Addict. Behav. 29, 89106.
  • Busnardo C., Tavares R. F., Resstel L. B., Elias L. L. and Correa F. M. (2010) Paraventricular nucleus modulates autonomic and neuroendocrine responses to acute restraint stress in rats. Auton. Neurosci. 158, 5157.
  • Campbell T., Lin S., DeVries C. and Lambert K. (2003) Coping strategies in male and female rats exposed to multiple stressors. Physiol. Behav. 78, 495504.
  • Choi D. C., Furay A. R., Evanson N. K., Ostrander M. M., Ulrich-Lai Y. M. and Herman J. P. (2007) Bed nucleus of the stria terminalis subregions differentially regulate hypothalamic-pituitary-adrenal axis activity: implications for the integration of limbic inputs. J. Neurosci. 27, 20252034.
  • Dallman M. F., Akana S. F., Jacobson L., Levin N., Cascio C. S. and Shinsako J. (1987) Characterization of corticosterone feedback regulation of ACTH secretion. Ann. N. Y. Acad. Sci. 512, 402414.
  • Diorio D., Viau V. and Meaney M. J. (1993) The role of the medial prefrontal cortex (cingulate gyrus) in the regulation of hypothalamic-pituitary-adrenal responses to stress. J. Neurosci. 13, 38393847.
  • Dluzen D. E. and Liu B. (2008) Gender differences in methamphetamine use and responses: a review. Gend. Med. 5, 2435.
  • Dunn J. D. and Orr S. E. (1984) Differential plasma corticosterone responses to hippocampal stimulation. Exp. Brain Res. 54, 16.
  • Fernandes G. A., Perks P., Cox N. K., Lightman S. L., Ingram C. D. and Shanks N. (2002) Habituation and cross-sensitization of stress-induced hypothalamic-pituitary-adrenal activity: effect of lesions in the paraventricular nucleus of the thalamus or bed nuclei of the stria terminalis. J. Neuroendocrinol. 14, 593602.
  • Fernández-Guasti A., Fiedler J. L., Herrera L. and Handa R. J. (2012) Sex, stress, and mood disorders: at the intersection of adrenal and gonadal hormones. Horm. Metab. Res. 44, 607618.
  • Figueiredo H. F., Dolgas C. M. and Herman J. P. (2002) Stress activation of cortex and hippocampus is modulated by sex and stage of estrus. Endocrinology 143, 25342540.
  • Franklin K. B. J. and Paxinos G. (2007) The mouse brain in stereotaxic coordinates, 3rd ed. Elsevier, Amsterdam.
  • Furay A. R., Bruestle A. E. and Herman J. P. (2008) The role of the forebrain glucocorticoid receptor in acute and chronic stress. Endocrinology 149, 54825490.
  • Giardino W. J., Pastor R., Anacker A. M., Spangler E., Cote D. M., Li J., Stenzel-Poore M. P., Phillips T. J. and Ryabinin A. E. (2011) Dissection of corticotropin-releasing factor system involvement in locomotor sensitivity to methamphetamine. Genes Brain Behav. 10, 7889.
  • Goel N., Plyler K. S., Daniels D. and Bale T. L. (2011) Androgenic influence on serotonergic activation of the HPA stress axis. Endocrinology 152, 20012010.
  • Gómez F., Lahmame A., de Kloet E. R. and Armario A. (1996) Hypothalamic-pituitary-adrenal response to chronic stress in five inbred rat strains: differential responses are mainly located at the adrenocortical level. Neuroendocrinology 63, 327337.
  • Handa R. J., Burgess L. H., Kerr J. E. and O'Keefe J. A. (1994a) Gonadal steroid hormone receptors and sex differences in the hypothalamo-pituitary-adrenal axis. Horm. Behav. 28, 464476.
  • Handa R. J., Nunley K. M., Lorens S. A., Louie J. P., McGivern R. F. and Bollnow M. R. (1994b) Androgen regulation of adrenocorticotropin and corticosterone secretion in the male rat following novelty and foot shock stressors. Physiol. Behav. 55, 117124.
  • Handa R. J., Weiser M. J. and Zuloaga D. G. (2009) A role for the androgen metabolite, 5alpha-androstane-3beta,17beta-diol, in modulating oestrogen receptor beta-mediated regulation of hormonal stress reactivity. J. Neuroendocrinol. 21, 351358.
  • Handelmann G. E. and Olton D. S. (1981) Spatial memory following damage to hippocampal CA3 pyramidal cells with kainic acid: impairment and recovery with preoperative training. Brain Res. 217, 4158.
  • Herman J. P., Patel P. D., Akil H. and Watson S. J. (1989) Localization and regulation of glucocorticoid and mineralocorticoid receptor messenger RNAs in the hippocampal formation of the rat. Mol. Endocrinol. 11, 18861894.
  • Herman J. P., Adams D. and Prewitt C. (1995) Regulatory changes in neuroendocrine stress-integrative circuitry produced by a variable stress paradigm. Neuroendocrinology 61, 180190.
  • Herman J. P., McKlveen J. M., Solomon M. B., Carvalho-Netto E. and Myers B. (2012) Neural regulation of the stress response: glucocorticoid feedback mechanisms. Braz. J. Med. Biol. Res. 45, 292298.
  • Hunsaker M. R., Tran G. T. and Kesner R. P. (2009) A behavioral analysis of the role of CA3 and CA1 subcortical efferents during classical fear conditioning. Behav. Neurosci. 123, 624630.
  • Isaacson R. L. and Lanthorn T. H. (1981) Hippocampal involvement in the pharmacologic induction of withdrawal-like behaviors. Fed Proc. 40, 15081512.
  • Jensen J. B., Jessop D. S., Harbuz M. S., Mørk A., Sánchez C. and Mikkelsen J. D. (1999) Acute and long-term treatments with the selective serotonin reuptake inhibitor citalopram modulate the HPA axis activity at different levels in male rats. J. Neuroendocrinol. 11, 465471.
  • de Kloet E. R. and Derijk R. (2004) Signaling pathways in brain involved in predisposition and pathogenesis of stress-related disease: genetic and kinetic factors affecting the MR/GR balance. Ann N Y Acad Sci. 1032, 1434.
  • de Kloet E. R., Joels M. and Holsboer F. (2005) Stress and the brain: from adaptation to disease. Neuroscience 6, 463475.
  • Kovács K. J. and Mezey E. (1987) Dexamethasone inhibits corticotropin-releasing factor gene expression in the rat paraventricular nucleus. Neuroendocrinology 46, 365368.
  • Kuo Y. M., Chen H. H., Shieh C. C., Chuang K. P., Cherng C. G. and Yu L. (2003) 4-Hydroxytamoxifen attenuates methamphetamine-induced nigrostriatal dopaminergic toxicity in intact and gonadetomized mice. J. Neurochem. 87, 14361443.
  • Leach L. S., Christensen H., Mackinnon A. J., Windsor T. D. and Butterworth P. (2008) Gender differences in depression and anxiety across the adult lifespan: the role of psychosocial mediators. Soc. Psychiatry Psychiatr. Epidemiol. 43, 983998.
  • Lowy M. T. (1990) MK-801 antagonizes methamphetamine-induced decreases in hippocampal and striatal corticosteroid receptors. Brain Res. 533, 348352.
  • McGivern R. F., Zuloaga D. G. and Handa R. J. (2009) Sex differences in stress-induced hyperthermia in rats: restraint versus confinement. Physiol. Behav. 98, 416420.
  • Nawata Y., Kitaichi K. and Yamamoto T. (2012) Increases of CRF in the amygdala are responsible for reinstatement of methamphetamine-seeking behavior induced by footshock. Pharmacol. Biochem. Behav. 101, 297302.
  • Nicosia N., Pacula R.L., Kilmer B., Lundberg R. and Chiesa J. (2009) The Economic Cost of Methamphetamine Use in the United States, MG-829-MPF/NIDA, 2009, p.169, online at:http://www.rand.org/pubs/monographs/MG829/ [accessed on 6 December 2013].
  • Palanza P. (2001) Animal models of anxiety and depression: how are females different? Neurosci. Biobehav. Rev. 25, 219233.
  • Plotsky P. M. (1987) Regulation of hypophysiotrophic factors mediating ACTH secretion. Ann. N. Y. Acad. Sci. 512, 205217.
  • Reichel C. M., Chan C. H., Ghee S. M. and See R. E. (2012) Sex differences in escalation of methamphetamine self-administration: cognitive and motivational consequences in rats. Psychopharmacology 223, 371380.
  • Reul J. M. and de Kloet E. R. (1985) Two receptor systems for corticosterone in rat brain: microdistribution and differential occupation. Endocrinology 117, 25052511.
  • Reul J. M. H. M., Van Den Bosch F. R. and De Kloet E. R. (1987) Differential response of type I and type II corticosteroid receptors to changes in plasma steroid level and circadian rhythmicity. Neuroendocrinology 45, 407412.
  • Robertson C. L. (1977) The regulation of vasopressin function in health and disease. Recent Prog. Horm. Res. 33, 333385.
  • Roth M. E. and Carroll M. E. (2004) Sex differences in the acquisition of IV methamphetamine self-administration and subsequent maintenance under a progressive ratio schedule in rats. Psychopharmacology 172, 443449.
  • Salomé N., Salchner P., Viltart O., Sequeira H., Wigger A., Landgraf R. and Singewald N. (2004) Neurobiological correlates of high (HAB) versus low anxiety-related behavior (LAB): differential Fos expression in HAB and LAB rats. Biol. Psychiatry 55, 715723.
  • Sawchenko P. E. and Swanson L. W. (1983) The organization of forebrain afferents to the paraventricular and supraoptic nuclei of the rat. J. Comp. Neurol. 218, 121144.
  • Sawchenko P. E., Arias C. A. and Mortrud M. T. (1993) Local tetrodotoxin blocks chronic stress effects on corticotropin-releasing factor and vasopressin messenger ribonucleic acids in hypophysiotropic neurons. J. Neuroendocrinol. 5, 341348.
  • Shoener J. A., Baig R. and Page K. C. (2006) Prenatal exposure to dexamethasone alters hippocampal drive on hypothalamic-pituitary-adrenal axis activity in adult male rats. Am. J. Physiol. Regul. Integr. Comp. Physiol. 290, 13661373.
  • Silverman A. J., Hoffman D. L. and Zimmerman E. A. (1981) The descending afferent connections of the paraventricular nucleus of the hypothalamus (PVN). Brain Res. Bull. 6, 4761.
  • Skorecki K. L., Brown D., Ercolani L. and Ausiello D. A. (1992) Molecular mechanisms of vasopressin action in the kidney, in Handbook of physiology, (Windhager E. E., ed.), pp. 11851218. Oxford UP, New York.
  • de Souza L. M. and Franci C. R. (2010) Differential immunoreactivity of glucocorticoid receptors and vasopressin in neurons of the anterior and medial parvocellular subdvisions of the hypothalamic paraventricular nucleus. Brain Res Bull. 82, 271278.
  • Substance Abuse and Mental Health Services Administration (SAMHSA) (2010) Gender differences among American Indian Treatment Admissions Aged 18 to 25. In: The TEDS Report, June 24, 2010. TEDS10-0624.
  • Substance Abuse and Mental Health Services Administration (SAMHSA). (2012) Results from the 2011 National Survey on Drug Use and Health: Summary of National Findings. In: NSDUH Series H-44, HHS Publication No. (SMA) 12-4713, Substance Abuse and Mental Health Services Administration, Rockville, MD.
  • Turner B. H. and Herkenham M. (1991) Thalamoamygdaloid projections in the rat: a test of the amygdala's role in sensory processing. J. Comp. Neurol. 313, 295325.
  • Uht R. M., McKelvy J. F., Harrison R. W. and Bohn M. C. (1988) Demonstration of glucocorticoid receptor-like immunoreactivity in glucocorticoid-sensitive vasopressin and corticotropin-releasing factor neurons in the hypothalamic paraventricular nucleus. J Neurosci Res. 19, 405–411, 468469.
  • Veening J. G., Bouwknecht J. A., Joosten H. J., Dederen P. J., Zethof T. J., Groenink L., van der Gugten J. and Olivier B. (2004) Stress-induced hyperthermia in the mouse: c-fos expression, corticosterone and temperature changes. Prog. Neuropsychopharmacol. Biol. Psychiatry 28, 699707.
  • Ventura-Silva A. P., Melo A., Ferreira A. C., Carvalho M. M., Campos F. L., Sousa N. and Pêgo J. M. (2013) Excitotoxic lesions in the central nucleus of the amygdala attenuate stress-induced anxiety behavior. Front. Behav. Neurosci. 7, 32.
  • Volpi S., Rabadan-Diehl C. and Aguilera G. (2004) Vasopressinergic regulation of the hypothalamic pituitary adrenal axis and stress adaptation. Stress 7, 7583.
  • Wang X. Y., Zhao M., Ghitza U. E., Li Y. Q. and Lu L. (2008) Stress impairs reconsolidation of drug memory via glucocorticoid receptors in the basolateral amygdala. J. Neurosci. 28, 56025610.
  • Wang H. N., Peng Y., Tan Q. R. et al. (2010) Quetiapine ameliorates anxiety-like behavior and cognitive impairments in stressed rats: implications for the treatment of posttraumatic stress disorder. Physiol. Res. 59, 263271.
  • Weiser M. J. and Handa R. J. (2009) Estrogen impairs glucocorticoid dependent negative feedback on the hypothalamic-pituitary-adrenal axis via estrogen receptor alpha within the hypothalamus. Neuroscience 159, 883895.
  • Williams M. T., Inman-Wood S. L., Morford L. L., McCrea A. E., Ruttle A. M., Moran M. S., Rock S. L. and Vorhees C. V. (2000) Preweaning treatment with methamphetamine induces increases in both corticosterone and ACTH in rats. Neurotoxicol. Teratol. 22, 751759.
  • Zavala J. K., Fernandez A. A. and Gosselink K. L. (2011) Female responses to acute and repeated restraint stress differ from those in males. Physiol. Behav. 104, 215221.
  • Zhu H., Mingler M. K., McBride M. L., Murphy A. J., Valenzuela D. M., Yancopoulos G. D., Williams M. T., Vorhees C. V. and Rothenberg M. E. (2010) Abnormal response to stress and impaired NPS-induced hyperlocomotion, anxiolytic effect and corticosterone increase in mice lacking NPSR1. Psychoneuroendocrinology 35, 11191132.
  • Zuloaga D. G., Morris J. A., Jordan C. L. and Breedlove S. M. (2008) Mice with the testicular feminization mutation demonstrate a role for androgen receptors in the regulation of anxiety-related behaviors and the hypothalamic-pituitary-adrenal axis. Horm. Behav. 54, 758766.
  • Zuloaga D. G., Poort J. E., Jordan C. L. and Breedlove S. M. (2011) Male rats with the testicular feminization mutation of the androgen receptor display elevated anxiety-related behavior and corticosterone response to mild stress. Horm. Behav. 60, 380388.
  • Zuloaga D. G., Siegel J. A., Agam M. and Raber J. (2013) Developmental methamphetamine exposure results in short- and long-term alterations in HPA axis-associated proteins. Dev. Neurosci. 35, 338346.