Olfactory bulbectomy increases food intake and hypothalamic neuropeptide Y in obesity-prone but not obesity-resistant rats

Obese individuals often suffer from depression. The olfactory bulbectomy (OBX) model is an animal model of depression that produces behavioral, physiological, and neurochemical alterations resembling clinical depression. The OBX model was employed to assess depression-related changes in food intake in obesity-prone, Osborne–Mendel (OM) rats and obesity-resistant, S5B/Pl rats. OBX increased food intake in OM rats beginning 7 days following surgery, however, OBX did not alter food intake in S5B/Pl rats at any time point. Fourteen days following surgery, OBX significantly increased locomotor activity (total lines crossed and rears) in the openfield test in OM and S5B/Pl rats. Fifteen days following surgery, prepro-neuropeptide Y (NPY) mRNA levels were significantly increased in the hypothalamus of bulbectomized OM rats and in the medial nucleus of the amygdala of bulbectomized OM and S5B/Pl rats. OBX decreased NPY Y2 receptor mRNA levels in the hypothalamus and medial nucleus of the amygdala in OM rats, while increasing NPY Y2 receptor mRNA levels in the medial nucleus of the amygdala of S5B/Pl rats. These data indicate that though both obesity-prone and obesity-resistant strains were susceptible to the locomotor effects of OBX, food intake and hypothalamic prepro-NPY mRNA were only increased in OM rats. Therefore, strain specific alterations in hypothalamic NPY may account for increased food intake in the obesity-prone rats following OBX, and suggests a potential mechanism to explain the comorbidity of obesity and depression.     

Herpes virus-mediated preproenkephalin gene transfer in the ventral striatum mimics behavioral changes produced by olfactory bulbectomy in rats

The syndrome of behavioral, physiological, and neurochemical changes caused by ablation of the olfactory bulbs (OBX) in rats serves as a reliable and well-validated model of depression. Previous experiments have demonstrated that OBX leads to increased expression of the preproenkephalin (ENK) gene in the olfactory tubercle (OT) portion of the ventral striatum in rats. The aim of the present experiments was to investigate the role of OBX-induced ENK overexpression in the OT in the behavioral abnormalities exhibited by bulbectomized rats. A recombinant herpes virus carrying human preproENK cDNA was used to manipulate ENK gene expression in the OT of bulbectomized and sham-operated rats. Motivational deficits were assessed by the sucrose preference test, and 'agitation-like' behaviors were measured with the novel open field and footshock-induced freezing tests. ENK gene transfer in sham-operated rats mimicked some of the effects of OBX; it decreased freezing behavior in response to mild footshock and produced behavioral activation in the open field. In another experiment, virally mediated ENK gene transfer into the OT of intact rats decreased footshock-induced freezing, and this effect was reversed by naltrexone administration. PreproENK gene transfer into the OT did not produce analgesic effects in the tail-flick test. No effects on freezing behavior were observed following preproENK gene transfer into the frontal cortex. An additional experiment revealed that naltrexone administration attenuated the OBX-induced abnormality in freezing behavior. The results indicate that overexpression of the preproENK gene in the ventral striatum may mediate the 'agitation-like' behavior exhibited by bulbectomized rats.     

Effects of chronic exercise and imipramine on mRNA for BDNF after olfactory bulbectomy in rat

We examined the effects of chronic activity wheel running and antidepressant treatment on brain-derived neurotrophic factor (BDNF) messenger RNA (mRNA) in multiple brain regions-hippocampal formation (HF), ventral tegmental area/substantia nigra (VTA/SN), nucleus accumbens (NAc), and piriform cortex (PFx)-after bilateral olfactory bulbectomy (OBX). Male, Long-Evans rats (n=72) underwent either sham or OBX surgery and were randomly divided into eight experimental groups in a 2 (sham vs. OBX) x 2 (sedentary vs. activity wheel)x2 (saline vs. imipramine) factorial design. Animals were killed after 21 days of treatment. Drug x exercise interaction effects were observed for HF (P=0.006-0.023) and VTA/SN (P=0.021); exercise increased BDNF mRNA in the saline treated animals but not in the imipramine treated animals. OBX did not affect BDNF mRNA in the HF or VTA/SN (P>0.05). BDNF mRNA levels in the PFx were not altered by exercise, drug, or OBX (P>0.05). These results suggest that the effect of exercise on BDNF mRNA extends beyond the HF to the mesolimbic ventral tegmental area and that the potentiation of BDNF mRNA by exercise and antidepressant pharmacotherapy, reported by other investigators, is time limited.     

Olfactory bulbectomy increases basal suprachiasmatic cyclic AMP levels in male rats.

For further characterization of the olfactory bulb's role in the medication of chronobiological phenomena, we examined basal cyclic- 3',5'-adenosine monophosphate (cAMP) levels in the suprachiasmatic nuclei (SCN) and lateral hippocampus (LHIP) following bilateral olfactory bulbectomy (OBX) to assess the effects of olfactory bulb removal on the biological clock. Two groups of adult Long-Evans rats underwent OBX or sham control surgery (SHAM). Eight weeks postoperative, the animals were decapitated at the time of maximal cAMP accumulation (circadian time 9-11 h), brains were removed, frozen in liquid nitrogen, and sectioned on a cryostat. Individual brain nuclei (SCN and LHIP) were microdissected using the Palkovits punch technique and analyzed by scintillation proximity assay for cAMP. We report a 83.6% increase in basal cAMP levels in the SCN following OBX (OBX = 63.7 pmol cAMP/mg protein, SHAM = 34.7 pmol cAMP/mg protein, p less than 0.01). No significant differences in LHIP cAMP levels were found. This specific increase in SCN cAMP, at the time of maximum cAMP accumulation, may give insight into the biochemical basis for altered activity levels following OBX.     

Leukemia inhibitory factor mRNA expression is upregulated in macrophages and olfactory receptor neurons after target ablation.

After target ablation by olfactory bulbectomy (OBX), the murine olfactory epithelium (OE) undergoes degenerative changes leading to apoptosis of olfactory receptor neurons (ORNs) followed by regenerative changes that include proliferation of progenitor cells leading to neurogenesis and ORN replacement. Macrophages recruited to the OE after OBX are involved in both the degenerative and regenerative processes. Relative quantitative RT-PCR was used to demonstrate that within hours of OBX, mRNAs encoding three key components in the leukemia inhibitory factor (LIF) signaling pathway, including LIF, LIF receptor (LIFR), and STAT3, as well as cyclin D1, a growth factor sensor indicative of progenitor cell transformation, were upregulated. These mRNAs reached peak levels of expression on or before day 3 post-OBX, coincident with the peak time for macrophage recruitment and progenitor cell proliferation. Cells expressing LIF mRNA in the OE of mice at 3 days post-OBX, the time point at which LIF mRNA expression peaked, were identified using non-isotopic in situ hybridization. LIF mRNA was localized in infiltrating macrophages; near-adjacent sections exhibited macrophages immunoreactive for F4/80, a marker for activated macrophages, in numbers commensurate with those expressing LIF mRNA. LIF mRNA was also localized in surviving ORNs, identified by their expression of olfactory marker protein (OMP) mRNA and protein in near-adjacent sections. Our data suggest that LIF functions as a mitogen originating from recruited macrophages through an intercellular signaling pathway that stimulates proliferation of progenitor cells leading to neurogenesis and regeneration, and as an intracellular survival factor for traumatized ORNs.     

Effects of olfactory bulbectomy on NMDA receptor density in the rat brain:

Olfactory bulbectomy (OBX) transects the glutamatergic efferents from the olfactory bulbs, and the changes of glutamatergic N-methyl-D-aspartate (NMDA) receptor-mediated function are though to be involved in the behavioral deficits seen in OBX rats. In the present study, irritability scores in OBX male Wistar rats were correlated with discrete regional effects on NMDA receptor function measured using a [3H] MK-801 binding assay. Irritability scores, measured before and for 2 weeks after OBX, showed a gradual increase in irritability after OBX. A reduction of the NMDA receptor density was observed in the cerebral cortex and amygdala 16 days after OBX, but not in the striatum, olfactory tubercle, entorhinal cortex, and hippocampus. These results demonstrate that OBX causes changes in the NMDA receptor system in certain brain regions and suggest that these changes may be responsible for the behavioral deficits of OBX rats.     

Effects of bilateral olfactory bulbectomy on N-methyl-D-aspartate receptor function: autoradiographic and behavioral studies in the rat

Rat bilateral olfactory bulbectomy (OBX) serves as a useful model in the study of depression and the mechanisms of action of antidepressant treatments. Considering the evidence of NMDA receptors involvement in depression, the present study was undertaken in order to investigate the time-course effects of OBX on the NMDA receptor function. Following bilateral olfactory bulbectomy, rats display an increase in locomotor activity and changes in other types of behavior in a novel environment. Autoradiographic experiments using the noncompetitive NMDA antagonist [(125)I]-iodo-MK-801 as the labeling agent showed that this increase in behavioral activities corresponds to a decrease in [(125)I]-iodo-MK-801 binding in a number of brain regions. In most regions, this reduction reached significance by the third week following OBX. However, in some cortical areas-a nucleus of the thalamus (AV) and one of the amygdala (LA)-this reduction was already significant in the first or second week following OBX and lasted throughout the 4 weeks of the study. We also compared the behavioral modifications induced by a challenge injection of MK-801 (0.2 mg/kg i.p.) in OBX and sham-operated rats. This challenge is known to induce hyperlocomotion and a number of stereotypies in naive rats. These effects were drastically reduced in OBX as compared to sham-operated rats. These data are consistent with the above-mentioned decrease in cerebral binding of MK-801 to NMDA receptors.     

CB1 modulation of hormone secretion, neuronal activation and mRNA expression following extracellular volume expansion

The endocannabinoid system includes important signaling molecules that are involved in several homeostatic and neuroendocrine functions. In the present study, we evaluated the effects of the type 1 cannabinoid (CB₁) receptor antagonist, rimonabant (10 mg/kg, p.o.), on hormone secretion, neuronal activation and mRNA expression in the hypothalamus following isotonic (I-) or hypertonic (H-) extracellular volume expansion (EVE). The total nitrate content in the PVN and SON was also assessed under the same experimental conditions. Our results showed that OT and AVP plasma concentrations were increased in response to H-EVE, while decreased AVP levels were found following I-EVE. Accordingly, both I- and H-EVE stimulated oxytocinergic neuronal activation, as evidenced by the increased number of c-Fos/OT double labeled neurons in the hypothalamus. The vasopressinergic cells of the PVN and SON, however, were only activated in response to H-EVE. Furthermore, increased amounts of both AVP and OT mRNAs were found in the hypothalamus following EVE. Pretreatment with rimonabant significantly potentiated hormone secretion and also vasopressinergic and oxytocinergic neuronal activation induced by EVE, although decreased AVP and OT mRNA expression was found in the hypothalami of rimonabant pretreated groups. In addition, the nitrate content in the PVN and SON was not altered in response to EVE or rimonabant pretreatment. Taken together, these results suggest that the CB₁ receptor may modulate several events that contribute to the development of appropriate responses to increased fluid volume and osmolality.     

Endocannabinoid modulation of amphetamine sensitization is disrupted in a rodent model of lesion‐induced dopamine dysregulation

We tested the hypothesis that increased dopaminergic sensitivity induced by olfactory bulbectomy is mediated by dysregulation of endocannabinoid signaling. Bilateral olfactory bulbectomy induces behavioral and neurobiological symptomatology related to increased dopaminergic sensitivity. Rats underwent olfactory bulbectomy or sham operations and were assessed 2 weeks later in two tests of hyperdopaminergic responsivity: locomotor response to novelty and locomotor sensitization to amphetamine. Amphetamine (1 mg/kg i.p.) was administered to rats once daily for 8 consecutive days to induce locomotor sensitization. URB597, an inhibitor of the anandamide hydrolyzing enzyme fatty‐acid amide hydrolase (FAAH), was administered daily (0.3 mg/kg i.p.) to sham and olfactory bulbectomized (OBX) rats to investigate the impact of FAAH inhibition on locomotor sensitization to amphetamine. Pharmacological specificity was evaluated with the CB₁ antagonist/inverse agonist rimonabant (1 mg/kg i.p). OBX rats exhibited heightened locomotor activity in response to exposure either to a novel open field or to amphetamine administration relative to sham‐operated rats. URB597 produced a CB₁‐mediated attenuation of amphetamine‐induced locomotor sensitization in sham‐operated rats. By contrast, URB597 failed to inhibit amphetamine sensitization in OBX rats. The present results demonstrate that enhanced endocannabinoid transmission attenuates development of amphetamine sensitization in intact animals but not in animals with OBX‐induced dopaminergic dysfunction. Our data collectively suggest that the endocannabinoid system is compromised in olfactory bulbectomized rats. Synapse 63:941–950, 2009. © 2009 Wiley‐Liss, Inc.     

Alterations in striatal and extrastriatal D-1 and D-2 dopamine receptors in the MPTP-treated common marmoset: An autoradiographic study

In adult common marmosets (Callithrix jacchus), MPTP (1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine) treatment induced almost total depletion of cells in the substantia nigra pars compacts (SNc) but partial cell loss in the ventral tegmental area (VTA). There was severe depletion of [³H]-mazindol binding to dopamine (DA) uptake sites in the caudate, putamen, and SNc. The loss of [³H]-mazindol binding in the nucleus accumbens (NAc) and olfactory tubercle (OT) was less marked. [³H]-mazindol binding in the body of caudate nucleus showed a small but significant recovery with increasing post-lesion survival times. The specific binding of [³H]-SCH 23390 to D-1 DA receptor sites was increased after MPTP treatment in all subregions of both caudate and putamen but was unaltered in the NAc and OT. Substantia nigra pars reticulata (SNr), frontal cortex, and medial segment of globus pallidus (GPm) all demonstrated moderate levels of [³H]-SCH 23390 binding in control animals, which were unaffected by MPTP treatment. Specific [³H]-spiperone binding to D-2 DA receptor sites was not altered by MPTP treatment in the subregions of caudate-putamen. Moderate levels of [³H]-spiperone binding were observed in control animals in the NAc, OT, SNc, and the lateral segment of globus pallidus (GPl). [³H]-spiperone binding in the SNc and OT was partially decreased in MPTP-treated animals. The changes in specific [³H]-spiperone and [³H]-SCH 23390 binding induced by MPTP-treatment did not alter with post-lesion survival times. These results demonstrate that MPTP treatment causes greater dopaminergic denervation of the caudate-putamen than in NAc/OT. This resulted in an increase in postsynaptic D-1 DA receptor sites in the caudate-putamen but not in the NAc/OT. Also, there appeared to be loss of presynaptic D-2 DA receptic sites in the SNc and OT. In the caudate-putamen, the loss of presynaptic D-2 DA receptor sites may have masked postsynaptic D-2 DA receptor upregulation. © 1993 Wiley-Liss, Inc.     

Cortical and nigral deafferentation and striatal cholinergic markers in the rat dorsal striatum: different effects on the expression of mRNAs encoding choline acetyltransferase and muscarinic m1 and m4 receptors

The regulation of the striatal m1 and m4 muscarinic receptor mRNA as well as the choline acetyltransferase (ChAT) mRNA expression by nigral dopaminergic and cortical glutamatergic afferent fibres was investigated using quantitative in situ hybridization histochemistry. The effects induced by a unilateral lesion of the medial forebrain bundle and a bilateral lesion of the sensorimotor (SM) cortex were analysed in the dorsal striatum 3 weeks after the lesions. Dopaminergic denervation of the striatum resulted in a marked decrease in the levels of m4 mRNA throughout the striatum, while the levels of muscarinic m1 mRNA and ChAT mRNA in cholinergic neurons were unaffected by the lesion. In contrast, following bilateral cortical ablation, the levels of the muscarinic m1 mRNA were significantly increased in the striatal projection area of the SM cortex, whereas the expression of m4 mRNA remained unchanged. Single cholinergic cell analysis by computer-assisted grain counting revealed a decreased labelling for ChAT mRNA per neuron following cortical ablation. However, in contrast to the topographical m1 mRNA changes, the decreased ChAT mRNA expression was evenly distributed within the striatum, suggesting an indirect cortical control upon striatal cholinergic interneurons. Altogether, these data suggest that dopaminergic nigral and glutamatergic cortical afferents modulate differentially cholinergic markers, at the pre- and post-synaptic levels. Beside the fact that nigral and cortical inputs exert an opposite control on cholinergic neurotransmission, our study further shows that this control involved different muscarinic receptor subtypes: the m4 and m1 receptors, respectively.     

Dynamic regulation of NGFI-A (zif268, egr1) gene expression in the striatum.

The expression of immediate-early genes of the fos/jun leucine zipper family can be regulated in striatal neurons by stimuli affecting the dopaminergic nigrostriatal system. The regulatory effects are gene specific, region specific, and striatal compartment specific. In the experiments reported here, we have explored the possibility that dopaminergic stimulation might also affect striatal expression of NGFI-A, a member of the zinc finger family of immediate-early genes. We treated healthy adult rats with amphetamine or cocaine and monitored the acute response of striatal neurons by in situ hybridization with oligonucleotide probes for NGFI-A mRNA. Both drugs evoked rapid, anatomically patterned increases in NGFI-A mRNA expression in the dorsal striatum (caudoputamen) and in the ventral striatum (nucleus accumbens, olfactory tubercle). The main response to each drug was in medium-sized neurons, known to be the projection neurons of the striatum. At every dose of amphetamine eliciting a response, the increased NGFI-A mRNA expression was preferentially concentrated in striosomes of the rostral caudoputamen, whereas cocaine at each dose given induced expression of NGFI-A mRNA in both striosomes and matrix at these striatal levels. The two indirect agonists evoked NGFI-A expression in both striatal compartments farther caudally, especially in the central and medial caudoputamen. Activation by both drugs was blocked by pretreatment with the D1-selective dopamine receptor antagonist SCH23390. These patterns of NGFI-A activation are remarkably similar to those found for Fos-like immunoreactivity following acute amphetamine and cocaine treatments, suggesting that coordinate activation of members of at least two immediate-early gene families occurs in the striatum following catecholaminergic stimulation. Such patterns of induction strongly support the view that the genomic responsiveness of the striosome and of the matrix compartments of the rostral striatum are distinct at the level of early-response gene expression. These findings raise the interesting possibility that some of the well-known effects of dopaminergic stimulation on neuropeptide and neurotransmitter expression in the striatum may reflect particular combinatorial patterns of immediate-early gene activation.     

Failure to detect collateral sprouting of mesolimbic dopaminergic neurons during early postnatal development

Using biochemical parameters the present study sought to assess the normal developmental pattern of the dopaminergic innervation of the olfactory tubercle (OT) and how it is affected by olfactory bulbectomy. In rats, the adult pattern of cellular organization is achieved in the OT gradually over the first 7 days after birth. On the other hand, tyrosine hydroxylase (TH) and [3H]dopamine ([3H]DA) uptake, while present at low levels, start to increase rapidly only after the first 7 days reaching adult levels by 40 and 20 days after birth, respectively. TH in dopamine (DA) cell bodies of A10 was already high, 40% of adult value, at birth, reached 150% by day 14 and decreased back to adult values by day 21 after birth. In 10-day-old rats, bulbectomy resulted, 30 days later, in an increase to 123% of control in TH activity and 137% in [3H]DA uptake within the OT. Comparable changes were found following bulbectomy in adults. However, bulbectomy in 1-day-old rats did not produce any significant changes 40 days later. The findings suggest that during postnatal growth TH activity is increased in DA cell bodies, preceding the changes in DA terminals of the OT, resembling the changes occurring during collateral sprouting in adults. In addition, changes indicative of collateral sprouting do not occur in response to deafferentation of the OT in 1-day-olds but do in 10-day-olds or older animals, a phenomenon probably related to a critical development period of the OT.     

Celecoxib delays cognitive decline in an animal model of neurodegeneration

Cyclooxygenase-2 (COX-2) is thought to play a role in the pathogenesis of various neurodegenerative disorders. However, clinical trials with COX-2 inhibitors have yielded contradictory results. In the present study we investigated whether COX-2 plays a role in the behavioral and cognitive impairments seen in olfactory bulbectomized rats. These impairments arise from neurodegenerative processes. First, we determined the time course of the OBX-induced behavioral (hyperactivity) and cognitive changes (fear memory) and how these correlate with changes in COX-2 mRNA expression in hippocampus. This experiment showed that the major impairments in behavior and cognition developed between Days 3 and 14 after OBX surgery, which correlated with changes in mRNA levels of COX-2, which increased at Days 7 and 14 after surgery but not anymore at day 28. In a subsequent experiment, rats were treated, starting two days before surgery, with the COX-2 inhibitor celecoxib (10mg/kg, dissolved in drinking water) for 4 weeks. OBX-induced hyperactivity in the open field was normalized after 2 weeks of celecoxib treatment, but not longer after 4 weeks. Celecoxib partly rescued fear learning and memory deficits without affecting spatial memory. The effects of celecoxib on fear memory lasted up to 1 week posttreatment, but disappeared thereafter. Our results show that COX-2 plays a limited role (both in magnitude and time) in the development of the OBX syndrome.     

Olfactory bulbectomy alters NMDA receptor levels in the rat prefrontal cortex

Olfactory bulbectomized (OBX) rats show a variety of behavioral and biochemical deficits that parallel human depression. We investigated the expression of glutamate receptor subtypes in cortical and subcortical brain regions following bilateral olfactory bulbectomy in adult rats. Quantitative receptor autoradiography using [(125)I]MK-801 (NMDA receptor), [(3)H]AMPA (AMPA receptor), and [(3)H]kainate (kainate receptor) was performed on brain sections at 1-5 weeks following olfactory bulbectomy. Our results show an elevation of NMDA receptors in the medial prefrontal cortex within 1 week following bulbectomy, which persisted up to at least 5 weeks post-bulbectomy. Neither kainate nor AMPA receptors were altered in any brain region examined. The potential significance of these results is discussed in light of experimental findings supporting a role for NMDA receptors in the mechanism of action of antidepressant drugs and the pathophysiology of major depression.     

Effects of prenatal cocaine exposure upon postnatal development of neostriatal dopaminergic function

Pregnant rats were injected twice daily with 20 mg/kg cocaine (or saline) from gestational day 10 to parturition. Brains from offspring were examined with quantitative receptor autoradiography [D1 receptor (D1R), D2 receptor (D2R) and dopamine transporter (DAT)] and quantitative in situ hybridization [D1R mRNA, D2R mRNA, preproenkephalin (PPE) mRNA] for markers of neostriatal dopaminergic function. Prenatal cocaine exposure did not alter postnatal development of striatal D1R sites, but D1R mRNA levels were reduced by a third at days 14 and 35. D2R sites were increased over control in lateral striatum by day 6, and remained elevated through postnatal day 35. Total D2R mRNA was increased over control in both medial and lateral striatum at 7 and 14 days but was equal to control at 35 days. Prenatal cocaine exposure increased DAT density at postnatal days 1 through 5, but reduced it at days 14 and 35; PPE mRNA expression was reduced at days 7, 14 and 35. Many of these results are similar to those found in experimental animals and humans following cocaine withdrawal. © 1994 Wiley-Liss, Inc.     

Changes in defensive behaviors following olfactory bulbectomy in male and female rats

The present study examined if olfactory bulbectomy (OBX) altered defensive behaviors on the elevated plus-maze and the open-field differently in male and female rats. Similar increases in defensive behaviors in male and female rats were observed in both tests following OBX. No significant correlations were detected between defensive behaviors and activity, supporting the hypothesis that some behavioral changes following OBX may be due to decreased defensive behaviors and not increased activity.