www.elsevier.com/locate/brainres

Brain Research 1046

Research report

Preliminary evidence for reduced social interactions in Chakragati

mutants modeling certain symptoms of schizophrenia

German Torresa, Beth A. Meederb,c, Brian H. Hallasa, Kenneth W. Grossc, Judith M. Horowitzb,c,*

aDepartment of Neuroscience, New York College of Osteopathic Medicine of New York Institute of Technology, Old Westbury, NY 11568, USAbClinical Neuroscience Laboratory, Medaille College, 18 Agassiz Circle, Buffalo, NY 14214, USA

cDepartment of Molecular and Cellular Biology, Roswell Park Cancer Institute, Buffalo, NY 14263, USA

Accepted 1 April 2005

Available online 10 May 2005

Abstract

Rodent models of schizophrenia provide powerful experimental tools for elucidating certain manifestations of the brain disease. The

chakragati (ckr) mouse mutant, for instance, reproduces aberrant neuroanatomical and behavioral phenotypes observed in the corresponding

human condition. To further investigate the utility of this mouse in the context of social behavior, we compared spontaneous behavioral

activity and social interactions recorded during the subjective night among wild-type, heterozygous, and homozygous ckr mice. We found

that both heterozygous and homozygous ckr animals failed to show appropriate norms of social behavior, including proximity, approach,

huddling, and anogenital investigation in response to novel conspecifics. We further found that the anatomical distribution, topography, and

connectivity of the neuropeptides oxytocin and vasopressin in the anterior hypothalamus did not differ among wild-type, heterozygous, or

homozygous ckr animals. These latter findings suggest that although oxytocin and vasopressin influence social behavior, connectivity of

such cells may not be phenotypically relevant for the observed social deficits seen in heterozygous and homozygous ckr mice. Collectively,

ckr mice and their heterozygote kin are valuable experimental tools for pre-clinical studies involving disruptions of social behavior (e.g.,

social withdrawal).

D 2005 Elsevier B.V. All rights reserved.

Theme: Disorders of the nervous system

Topic: Neuropsychiatric disorders

Keywords: Behavioral activity; Brain; Circling behavior; Ckr mice; Oxytocin; Social withdrawal; Vasopressin

1. Introduction

Schizophrenia is a brain disease with variable deficit

manifestations in behavior. Deficit symptoms (also com-

monly referred to as negative symptoms) include blunted

affect, social ambivalence, and social withdrawal [2,17]. The

observation that schizophrenic patients have deficits in social

interaction suggests that certain neural circuits processing

social information have been pathologically modified by the

0006-8993/$ - see front matter D 2005 Elsevier B.V. All rights reserved.

doi:10.1016/j.brainres.2005.04.015

* Corresponding author. Clinical Neuroscience Laboratory, Medaille

College, 18 Agassiz Circle, Buffalo, NY 14214, USA. Fax: +1 716 884

0291.

E-mail address: jhorowitz@medaille.edu (J.M. Horowitz).

disease. To achieve a better understanding of brain–behavior

relationships, particularly those that are symptomatic of

schizophrenia, genetically engineered mouse mutants might

be relevant for investigating aberrant social behavior.

Currently there are several genetic animal models of schizo-

phrenia that reproduce certain aspects of the human disease

phenotype [3,13,19,20,22]. The chakragati (ckr) mouse, for

instance, shows behavioral abnormalities, including novelty-

driven hyperactivity that is manifested as increased circling

activity in the open field. Importantly, this schizophrenia-

related behavior is attenuated by treatment with the anti-

psychotics clozapine and olanzapine [19]. Further, ckr mice

show asymmetric elevation of dopamine D2-like receptors in

the striatum, and high-resolution structural MRIs of these

(2005) 180 – 186

G. Torres et al. / Brain Research 1046 (2005) 180–186 181

mutants indicate selective enlargement of the lateral ven-

tricles, changes that are reminiscent of schizophrenia [14,20].

We have previously reported that the phenotype of the

ckr mouse is transmitted as an autosomal recessive trait,

suggesting that it may result from a loss of function at a

particular locus (e.g., D16Ros1) of chromosome 16 [14,16].

Determination of the disrupted gene(s) responsible for this

phenotype may offer insight into the control of complex

motor behaviors. Thus, ckr mutants are not only amenable

to pre-clinical interventional trials but are also useful to

characterize the behavioral effects of experimentally

induced mutations. Against this background and to further

investigate the behavioral significance of the Ren-2d renin

gene insertion in the mouse genome, male ckr mice were

paired with unfamiliar (female) conspecifics and several

behavioral features of social behavior were videotaped

during the subjective night. In addition, as the neuro-

hypophyseal neuropeptides oxytocin (OT) and arginine

vasopressin (AVP) have been shown to influence a number

of forms of social behavior, including affiliation and

reproduction [1,24], we mapped the anatomical distribution

of OT and AVP nerve cells in brains of ckr (ckr/ckr),

heterozygous (+/ckr), and wild-type (+/+; C57BL/10Rospd�C3H/HeRos) mice. In general our data suggest that the ckr

mouse mutant mimics certain behavioral symptoms of

schizophrenia, namely deficits in social interactions.

2. Materials and methods

2.1. Animals and experimental design

The ckr mouse was generated as described previously

[14]. All transgenic mice used for the experiments below

were adult (6–9 months; 30–40 g) male and female F2animals of the mixed genetic background of BCF1 (C57BL/

10Rospd � C3H/HeRos). Homozygous, heterozygous, and

wild-type adult mice were kept in (same-sex/same-geno-

type) groups of 3–4/cage and maintained on a light–dark

cycle of 12:12 h (lights on at 0700) with free access to food

and water. Mice (from different litters) were never handled

or isolated before the social interaction test paradigm (see

below). Classification of genotype for both ckr and

heterozygous mice was conducted by restriction fragment-

length polymorphism analysis of biopsied tail DNA taken

during the first week of postnatal life [14]. All behavioral

procedures were carried out in accordance with the NIH

Guide for the Care and Use of Laboratory Animals, and with

approval from the Roswell Park Cancer Institute IACUC.

All efforts were made to minimize animal stress and to

reduce the number of mice used for these experiments.

2.2. Behavioral testing: general behavioral activity

Adult wild-type, heterozygous, and homozygous ckr

mice housed in their respective home-cages were videotaped

(under an infrared light emitting wavelengths greater than

680 nm) for 60 min during the subjective night (subjective

night denotes the onset of daily activity). In brief, sponta-

neous behavioral activity was recorded (4 h after subjective

night onset) on a Sony Model TRV 900 videotape recorder

equipped with a standard 35 mm Sony mini-DV tape.

Overall behavioral activity, including circling behavior, was

digitized for 1 min and the number of complete, full (left or

right) 360- circles was scored by two investigators with no

knowledge of the genotype identity. In addition, the

percentage of time spent on walking, rearing, or time spent

on stationary activities (e.g., grooming, sniffing, or nibbling)

was quantified from the video recordings (for further

methodological details see Ref. [19]).

2.3. Behavioral testing: social interactions in neutral cages

To assess social behavior between two mice of the

opposite sex in a neutral environment, first we videotaped

genetically identical mice that had been housed separately

by sex and then placed in a home-cage for one night. Testing

began when a stimulus female mouse was introduced into the

neutral environment of a male animal habituated to that

environment 30 min prior to the social confrontation. The

neutral home-cage was a rectangular Plexiglas cage (30 cm�15 cm � 20 cm) with food and water, containing the above

infrared video camera. Recorded spontaneous social inter-

actions were digitized for each mouse using a Peak Motus

Program. This was achieved by marking several constant

points on every animal recorded: the nose, occiput, left and

right shoulders, left and right hips, and the base of the tail.

The aforementioned coordinates were followed for 3 s to

generate a spatial diagram of motor activities by plotting the

range of circular motion angles (for further methodological

details see Ref. [6]). Second, duration of social interactions

(i.e., proximity, approach, and huddling) and investigative

behaviors (i.e., olfactory exploration and anogenital inves-

tigation) were scored and quantified from the video record-

ings for a 60-min period by investigators with no knowledge

of the mouse genotype(s). Recordings began 4 h after the

onset of the subjective night. After the testing procedures,

animals (N = 4 mice/genotype) were returned to their

original, respective home-cages.

2.4. Olfactory test

To determine olfactory performance, wild-type, hetero-

zygous, and homozygous ckr animals (N = 4 mice/

genotype) were placed individually in a neutral cage 4 h

after the onset of the subjective night. An investigator blind

to the genotype of the mouse tested hid a miniature Oreo

cookie in the cage bedding and immediately recorded the

latency (in seconds) of each individual mouse to locate and

retrieve the cookie. Two successive trials were performed

for each mouse genotype under an infrared light emitting

wavelengths greater than 680 nm.

Fig. 1. Spontaneous behavioral activity in mice. Stereotypical locomotor

parameters recorded during the subjective night in the home-cage revealed

increased rotational activity in homozygous ckr mice relative to wild-type

and heterozygous animals. *P � 0.05. It is unlikely that specific behavioral

abnormalities in ckr mice are a reflection of general health effects, as these

animals appear grossly normal and healthy. Data depict means T SEM

during a cumulative 60-min video-imaging tracking session. N = 5 male/

female mice/genotype.

G. Torres et al. / Brain Research 1046 (2005) 180–186182

2.5. Immunocytochemistry for neuropeptides

Mice used for the immunocytochemical experiments

were deeply anesthetized with 4% isoflurane and rapidly

decapitated. Brains were collected and placed in 4%

paraformaldehyde in Na2PO4 buffer solution (pH 7.2) for

5 days at 4 -C and then stored overnight in 20% sucrose

(dissolved in 0.01 M sodium phosphate buffer). Prior to

sectioning, brains were frozen with dry ice, mounted on a

sliding microtome, and cut into 50 Am coronal sections

from rostral striata to cerebella. Cut brain sections were

collected in a cold cryoprotectant solution (0.05 M sodium

phosphate buffer pH 7.3, 30% ethylene glycol, and 20%

glycerol) and stored at �20 -C until prepared for standard

immunocytochemical procedures. To unmask target epito-

pes in brain material, free-floating sections were first

washed with 0.5 M potassium-phosphate buffer solution

(KPBS) and then were incubated in a sodium citrate buffer

solution (10 mM, pH 9.0; Sigma, St. Louis, MO) for 30 min

in a water-bath set to 80 -C [7,9]. Following the above

antigen retrieval, brain sections were incubated for 48 h at 4

-C with rabbit polyclonal IgGs raised against OT and AVP

(both antibodies obtained from Chemicon International,

Temecula, CA), diluted 1:1000 in KPBS with 1% normal

goat serum and 0.3% Triton X-100. Next, brain sections

were rinsed in KPBS and incubated with a secondary

antibody biotinylated goat anti-rabbit IgG (1:1500 dilution;

Vector Laboratories, Burlingame, CA) for 60 min. Sub-

sequently, brain sections were incubated at room temperature

with an avidin–biotin–peroxidase complex (Vectastain

ABC Elite Kit; Vector laboratories) for 60 min. After

several rinses in KPBS and 0.05 M acetate-imidazole buffer,

brain sections were developed in a mixture containing tris-

imidazole buffer, nickel-sulfate, and the chromagen 3,3-

diaminobenzidine tetrahydrochloride (DAB) along with 1%

H2O2. Following this developing phase, brain sections were

again washed in KPBS, mounted onto gelatin-chrome-alum-

coated slides, allowed to dry overnight, dehydrated through

graded alcohols, cleared in xylenes, and cover-slipped with

DPX mountant (Electron Microscopy Sciences, Ft. Wash-

ington, PA). To help identify relevant anatomical structures,

brain sections were counterstained with Neutral Red.

Specificity for the above antibodies was instituted in the

form of negative controls (i.e., replacing the primary

antibody with serum). Visualization of OT- and AVP-

positive neurons was accomplished with bright-field micro-

scopy using an Olympus microscope at magnifications of

�10, �20, and �40.

2.6. Data analysis

Behavioral data are presented as means T SEM. To test

for significant differences between group means, data were

analyzed by ANOVA followed by Newman–Keuls post hoc

test comparisons. Sex was not used as a main effect factor in

the ANOVAs as no apparent differences in circling behavior

or hypothalamic cell number were noted between male or

female mice of a given genotype. Statistically significant

differences were defined as P � 0.05. Neuroanatomical

data are also presented as means T SEM of labeled

neurons. The number of OT- and AVO-positive cells was

estimated for hypothalamic sections obtained from wild-

type, heterozygous, and homozygous ckr mice. Using a

modified version of the fractionator method [23], OT- and

AVP-labeled neurons were counted (per 1 mm2) in the

hypothalamus of every 4th serial brain section through the

parvocellular section of the mouse hypothalamus on coded

slides (for a total of eight brain sections). Cell counts were

analyzed by ANOVA followed by Newman–Keuls post

hoc test comparisons with statistical significance defined as

P � 0.05.

3. Results

3.1. General behavioral activity

Wild-type and heterozygous mice videotaped for 1 h

during the subjective night showed relatively low circling

behavior as defined by complete (360-) rotational turns in

place. In contrast, homozygous ckr mice displayed a robust,

aberrant circling behavior phenotype throughout the entire

testing session (Fig. 1). As previously described [19], male

and female ckr animals voluntarily engaged in individual

turning rates ranging from 50 to 70 full body turns/min (no

differences in turning rates were apparent between sexes).

After 1 h of testing, ckr mouse mutants (of both sexes) had

primarily engaged in spinning using the hind limbs as a

pivot to reach bouts of circling reaching near 1200 full body

turns (Fig. 2). Further, circling mutants displayed very little

Fig. 2. Time-course behavioral observations. Data depict means T SEM for

rotational activity displayed by ckr mice during the subjective night in the

home-cage. Note the persistent aberrant activity of homozygous mutants

during each video-imaging tracking session. In contrast, heterozygous mice

did not show a circling behavior phenotype. N = 5 male/female mice/

genotype. *P � 0.05 when compared with wild-type and heterozygous

cohorts.

Fig. 3. Digitized video-images depicting mouse social interactions in

neutral cages. Male (green line) and female (purple line) wild-type animals

showed several proximity interactions during each of four 3 s digitized

trials. In contrast, spatial proximity interactions between male and female

conspecifics are decreased in heterozygous and homozygous ckr mice.

Note the range of circular motions shown by homozygous cohorts.

Although circling behavior was increased in the ckr couple, it is possible

that their decreased duration of social contacts was a consequence of

increased behavioral activity. However, the time that ckr mutants spent

separated from each other was not significantly greater than that of

heterozygous mice (see Table 1).

G. Torres et al. / Brain Research 1046 (2005) 180–186 183

grooming or rearing activity when compared with wild-type

and heterozygous mice of both genders. Thus, hyperactivity

as manifested by increased circling activity in the home-

cage is a predominant behavioral feature of the ckr mouse.

This observation further supports the occurrence of signifi-

cant (F2,11 = 7.84, P � 0.05) genotype-dependent differ-

ences in spontaneous behavioral activity. Of potential

interest, homozygous mutants often attempted to copulate

during the recorded session suggesting some type of

hypersexual behavioral trait (data not shown). In this regard,

hypersexuality is also present in some but not all cases of

schizophrenia [21]. Finally, it should be noted that there is

no indication of hearing loss or degeneration of the bony

and membranous labyrinths of the ckr mouse inner ear, thus

supporting the hypothesis that a central, rather than a

vestibular, derangement is causing the circling phenotype

[14].

3.2. Social interactions

We used measurements for proximity, approach, hud-

dling, and anogenital investigation of two animals (male and

female), derived from video images, to quantify the role of

genotype in social interactions (Fig. 3). We define social

interaction as any detectable change in the behavior of one

animal caused by another [10]. First, male wild-type animals

when presented with a stimulus conspecific female showed

vigorous investigative behavior, including close following

inspection (proximity and approach) and bouts of intense

G. Torres et al. / Brain Research 1046 (2005) 180–186184

anogenital sniffing. Relative to males, wild-type females

displayed far lower levels of baseline investigatory behav-

ior. Nevertheless, they showed conspicuous proximity and

approach interactions with resident males as detected by the

amount of time spent in intentional social interactions. Side-

by-side posture, referred to as huddling, was also measured

in wild-type mice with significant amounts of physical

contact spent in this affiliative behavior. In contrast, animals

carrying either a single (i.e., heterozygous) or double (i.e.,

homozygous) copy of a transgene insertion showed sig-

nificant (P � 0.05) differences in the above social

interactions. For example, when confronted with unfamiliar

conspecifics, heterozygous mice exhibited little or no

olfactory exploration or huddling behavior during the

defined testing session. Hence, a characteristic decline in

the total time spent in intentional social interactions was

recorded for this mouse genotype (Table 1).

Disruptions in social interactions were also recorded in

homozygous ckr mice when presented with a stimulus

conspecific during the subjective night. In general, prox-

imity, approach, and huddling behaviors as well as

anogenital sniffing were significantly restricted in ckr mice.

As a consequence, this particular mouse genotype also

showed a characteristic decline in the total amount of time

spent in intentional social interactions (Table 1). Thus, both

heterozygous and homozygous mice showed comparable

qualitative deficits in social interaction parameters, but only

one genotype, the homozygous ckr, displayed an aberrant

circling phenotype. This finding suggests that decreased

duration of social interactions by homozygous ckr mice is

not a consequence of increased behavioral activity per se.

Rather it appears that onset of social deficits is the result of

carrying single or double copies of the Ren-2d renin gene.

Further our findings suggest that social behavior, partic-

ularly the natural tendency of mice to intensely investigate

conspecifics, has a neural basis distinct from that of circling

behavior. Finally it should be noted that we failed to detect

impairments in olfactory function in heterozygous or

homozygous animals as both genotypes, when compared

Table 1

Spontaneous social interactions in mice recorded during a 60-min

observational trial

Genotype Proximity, approach,

and huddling

Olfactory exploration

and anogenital investigation

Wild-type 5 min 18 s T 38 s* 3 min 42 s T 14 s*

Heterozygous 1 min 1 s T 26 s 1 min 3 s T 41 s

Homozygous 0 min 23 s T 10 s 1 min 5 s T 5 s

Under this social recognition paradigm, heterozygous and homozygous ckr

mice recognize non-social signals. Each value is the amount of time spent

in social interactions (means T SEM). Overall one-way ANOVA for social

encounters yielded *P � 0.05 and *P � 0.05 for Newman–Keuls post hoc

test comparisons between wild-type and heterozygous/homozygous ckr

animals. The number of social contacts between heterozygous and

homozygous ckr cohorts did not differ significantly (P � 0.05). Spatial

memory, olfactory memory, and/or habituation have not yet been tested.

with wild-type mice, showed equal performance in locating

buried food (Oreo cookies) in two successive trials. In this

regard, means T SEM in retrieval time (seconds) for wild-

type mice = 4.25 T 0.62; for heterozygous cohorts = 4.0 T0.40; and for homozygous ckr animals = 3.5 T 0.85 (F2,11 =

0.63, P � 0.05 as detected by one-way ANOVA). These

data indicate that although levels of social interaction differ

between genotypes, olfactory capabilities do not.

3.3. OT and AVP connectivity in the ckr mouse brain

Several studies suggest that social behavior, notwith-

standing its complexity, has a neural basis involving the

neuropeptides OT and AVP [1,8]. To determine whether the

gross cytological and connectional characteristics of OT and

AVP had been modified by the insertion of the Ren-2d renin

gene, we evaluated OT and AVP neuronal populations

ordinarily present in the medial parvocellular division of the

paraventricular nucleus (PVNp) of the hypothalamus of

wild-type, heterozygous, and homozygous ckr mouse

brains. Representative hypothalamic coronal (50 Am)

sections of mature wild-type animals contained numerous

OT and AVP immunoreactive (IR) neurons. These cells

tended to be grouped into clusters of the PVNp and showed

both fine caliber fibers and dendrites (Fig. 4). Like the OT-

and AVP-containing neurons of wild-type control hypothal-

ami (means T SEM = 40.3 T 1.19; 98.4 T 3.3, respectively),

the neuropeptide population of heterozygous (39.5 T 1.5;

99.0 T 3.8, respectively) and homozygous (40.1 T 0.8;

101.1 T 1.8, respectively) mice showed oval or round cells

organized into large, well-defined clusters of oxytocinergic

and vasopressinergic components (F2,11 = 0.1, P � 0.05 as

detected by one-way ANOVA for OT and F2,11 = 0.2, P �0.05 as detected by one-way ANOVA for AVP as well).

Immunoreactive fibers of very fine caliber ramified exten-

sively around the cell bodies of origin and in their

immediate vicinity, projecting extensively to distal aspects

of the anterior hypothalamus. Although it was difficult to

follow the dendrites for any significant length in 50 Amsections, the proximal dendritic structure of the mutant mice

agreed well with our observations from the wild-type brain

material and was characteristic of the isodendritic form

previously ascribed to numerous hypothalamic peptidergic

neurons [15]. Further, OT and AVP cells of the supraoptic

nucleus (SON) as well as cells of the accessory nucleus of

the magnocellular neurosecretory system (i.e., nucleus

circularis) recapitulated the anatomical features of wild-type

hypothalamus. These observations demonstrate that anterior

hypothalami and peptidergic cell types of heterozygous and

homozygous brains show many of the basic morphological

and connectivity features of the normal, mature mouse

brain. Thus conservation of neurohistologic OT and AVP

type is a general feature of all three genotypes and suggests

that deficits in social behavior amply recorded in hetero-

zygous and homozygous ckr mice do not appear to be

related to differences in hypothalamic morphology, cell

Fig. 4. OT and AVP (pseudo-color) neurons of the mouse hypothalamus. Numerous medium-sized and small neuropeptide cells are seen in wild-type,

heterozygous, and homozygous ckr brain material. N = 4 male mice/genotype. No gross differences in the number, morphology, or distribution of OT and AVP

neurons are apparent in the PVNp of each genotype. Native receptor distribution for the above neuropeptides has not yet been performed. Hypothalamic

sections depicted represent bregma �0.50 mm to bregma �0.70 mm of the comparative cytoarchitectonic atlas of the C57BL/6 and 129/Sv mouse brains [4].

Magnification �40.

G. Torres et al. / Brain Research 1046 (2005) 180–186 185

connectivity, or overall gross chemical neural circuitry.

Finally, no sex-dependent differences in any of the above

anatomical parameters were noted between male and female

mice of a given genotype.

4. Discussion

We have provided additional information regarding the

use of the ckr mouse mutant as a model for studying certain

manifestations of schizophrenia. Deficits and alterations in

social behavior including emotional withdrawal, blunted

affect, apathy, and disinterest in social familiarity are highly

pernicious symptoms of schizophrenia and other psychiatric

disorders [1]. Although such symptoms cannot always

easily be translated into behaviors that can be defined and

operationalized in animal models, they can nevertheless be

broken down into simple phenotypes that can be tested in

both human populations and in animal studies. The

behavioral aspects of social approach, recognition, and

conspecific interaction are relatively simple observational

tests often conducted in clinical presentations and animal

experiments [12,18]. In this regard, mice have proven

extremely valuable for understanding the basic neurobio-

logical mechanisms underlying social information [1,24].

Indeed, the natural tendency of rodents to intensely

investigate novel conspecifics highlights the amenability

of mice to examine social behaviors deemed abnormal in

human patients. In this report, we have characterized social

interactions in both male and female ckr mice during

subjective night pairing. In general, we show that hetero-

zygous and homozygous ckr animals exhibited a character-

istic decline in the time spent investigating novel

conspecifics. In contrast, social behavior of wild-type mice

(C57BL/10Rospd � C3H/HeRos; the genetic background

into which the Ren-2d renin gene was introduced) was

stereotypical in time spent investigating unfamiliar stimulus

animals. These data demonstrate significant genotype-

dependent differences in social behavior. It should be noted

that there is no over-expression of renin in the ckr mouse

line, therefore precluding the possibility that excessive

amounts of aspartyl protease are responsible for the above

social deficits [14]. Thus, it is conceivable that the transgene

(i.e., Ren-2d renin gene) insertion responsible for the

apparent lack of social recognition disrupted multiple

(unknown) transcripts within mouse chromosome 16.

Our studies indicate that ckr mice are valuable animals

for anti-psychotic drug testing, and also for high-resolution

magnetic resonance (MR) imaging studies involving brain

lesions [19,20]. The current findings extend the utility of the

above transgenic mice by demonstrating deficits in social

interactions as well. Thus, it seems that ckr mice and their

heterozygote kin have obvious behavioral and brain

abnormalities relevant to schizophrenia. The fact that

heterozygous animals showed equivalent deficits in social

behavior as homozygous ckr mice is indicative that both

mutants lack social recognition traits; with the heterozygote

kin having a milder circling phenotype. This is of significant

clinical interest as relatives of schizophrenics who do not

express the full spectrum of negative and positive symptoms

nevertheless manifest a number of social deficits as well [5].

Thus disorder symptoms (e.g., apathy or anhedonia) often

described in ‘‘schizophrenic’’ families seem to depend, in

part, on continuum gradients that are not only quantitative

but also qualitatively different from normal experience.

Heterozygote kin are therefore ideal animals for decon-

structing the molecular events leading to some of the

behavioral abnormalities that are seen in homozygous ckr

mice. Indeed, this ‘‘family-up approach’’, starting from a

non-afflicted kin and then progressing to deduce core

psychotic behaviors in afflicted individuals, has already

provided evidence of several disease mechanisms under-

lying the brain disorder [11]. This insight could have not

been gained from the study of schizophrenic patients alone.

G. Torres et al. / Brain Research 1046 (2005) 180–186186

Behavioral traits such as social interactions appear to be

quantitative traits that are normally distributed in both

human and rodent populations. Like humans, rodents seek

social contact; and because mice share important physio-

logical, anatomical, and genomic similarities with humans,

it is expected that both species contain core cellular systems

regulating social behavior. In these mammalian species,

most of the findings relating cellular systems to social

behaviors have focused on the nine-amino acid peptides OT

and AVP [8]. Here, we have found that the distribution

pattern of the above cell-containing molecules in homo-

zygous ckr and heterozygous mice was highly reproducible

and similar to the reported distribution of OT- and AVP-IR

in the mature, wild-type mouse brain. We interpret this

finding to mean that genotype-dependent differences in

social behavior are not due to differences in hypothalamic

neuropeptide labeling and/or topographical distribution. The

fact that we failed to detect genotype-dependent differences

in OT- and AVP-IR among animals showing striking

differences in social recognition, approach, and avoidance

raises the question as to the source of such differences. We

have previously reported that the phenotype of the ckr

mouse is transmitted as an autosomal recessive trait,

suggesting that it may result from a loss of function at a

particular locus (e.g., D16Ros1) of chromosome 16 [14,16].

Unfortunately, it is not yet clear what gene(s) have been

deleted or disrupted by the insertion of the Ren-2d renin

gene. Currently, we are attempting to identify the genetic

lesion responsible for the abnormal social behavior in ckr

mice. This may ultimately lead to the discovery of genes,

proteins, and neural circuits involved in schizophrenia

pathogenesis.

Acknowledgments

This study was supported by an NIH grant (#1R15MH

64513-01A1) to J.M.H., and in part by a grant (#CA-76561)

and an Institute Comprehensive Cancer Center Support

Grant (#CA-16056) from the National Cancer Institute,

Bethesda, MD. The authors wish to thank Riya Jose for her

technical assistance and Holly Johnson (New Media

Institute) for her assistance with digital imaging.

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