Dermorphin-SAP / MOR-SAP References

Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

69 entries found for : it-12

The role of the patch compartment of striatum in reward-driven behaviors

Ahn J, Christy DJ, Horner K (2021) The role of the patch compartment of striatum in reward-driven behaviors. Neuroscience 2021 Abstracts P747/06. Society for Neuroscience, Virtual.

Summary: The striatum is a neural structure that plays a critical role in cognitive functions, behavioral decision-making, and reward generation. The striatum exhibits a heterogeneous composition, containing neurons belonging to the patch compartment—which is thought to be involved in habitual reward-related behaviors—surrounded by neurons belonging to the matrix compartment—which is thought to be involved in adaptive motor control. Additionally, the striatum is further subdivided into the dorsolateral striatum (DLS) and the dorsomedial striatum (DMS), each with their own patch and matrix compartments. The DMS has been associated with goal-oriented behavior seen during the initial stages of addiction. Conversely, the DLS has been associated with habitual behaviors seen during late-stage addictive behaviors that are inflexible. It is thought that drug addiction is initially mediated by the DMS before DLS activity becomes predominant. Previously, it has been shown that the patch compartment of the DLS is necessary for development of habitual behavior, but the role of the patch compartment of the DMS is less clear. Our study intends to demonstrate that selective ablation of DMS patch compartment neurons will result in a negative impact on the initial development of reward-driven behaviors during the early stages of drug addiction. Since patch compartment neurons express a high level of mu opioid receptors compared to the surrounding matrix, we used dermorphin-saporin, a toxin that selectively destroys mu opioid receptor-containing neurons to target patch compartment neurons in the DMS and DLS for ablation. Following infusion in the DMS or DLS with dermorphin-saporin (17 ng/μl) or vehicle, rats were trained to self-administer cocaine (0.4 mg/kg/infusion) on progressive ratio schedule of reinforcement, starting with fixed ratio of 1 and ending with a fixed ratio of 5. Ablation of the patch compartment altered the level of responding for cocaine as the schedule of reinforcement became progressively labor-intensive. These data suggest that the patch compartment contributes to reward-driven behaviors.

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Lesions of the patch compartment of dorsolateral striatum disrupt stimulus-response learning.

Jenrette TA, Logue JB, Horner KA (2019) Lesions of the patch compartment of dorsolateral striatum disrupt stimulus-response learning. Neuroscience 415:161-172. doi: 10.1016/j.neuroscience.2019.07.033

Objective: To investigate whether enhanced activation of the patch compartment contributes to habitual behavior.

Summary: The dorsolateral patch compartment may mediate habit formation by altering information flow through basal ganglia circuits.

Usage: A volume of 2 ul of Dermorphin-SAP (17 ng/ul or an equivalent amount of unconjugated SAP (as a control) was infused bilaterally, at a rate of 0.5 ul/min.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Inflammatory mediators of opioid tolerance: Implications for dependency and addiction.

Eidson LN, Murphy AZ (2019) Inflammatory mediators of opioid tolerance: Implications for dependency and addiction. Peptides 115:51-58. doi: 10.1016/j.peptides.2019.01.003

Objective: To determine what mediates opioid tolerance and alterations in glutamate homeostasis.

Summary: Site-specific lesions of PAG MOR-containing neurons using Dermorphin-SAP significantly reduce the antinociceptive effects of systemic morphine suggesting that PAG MOR is critical for morphine action.

Usage: Rats were injected with 3 pmol of Dermorphin-SAP (Cat. #IT-12) into the PAG. Blank-SAP (Cat. #IT-21) was used as a control.

See: Loyd DR et al. Sex differences in micro-opioid receptor expression in the rat midbrain periaqueductal gray are essential for eliciting sex differences in morphine analgesia. J Neurosci 28:14007-14017, 2008.

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Selective ablation of striatal striosomes produces the deregulation of dopamine nigrostriatal pathway

Shumilov K, Real MÁ, Valderrama-Carvajal A, Rivera A (2018) Selective ablation of striatal striosomes produces the deregulation of dopamine nigrostriatal pathway. PLoS One 13:e0203135. doi: 10.1371/journal.pone.0203135

Objective: To increase knowledge in the role of the striosomal projection onto the dopamine neurons of the SNc and its impact on the nigrostriatal dopamine pathway.

Summary: Results highlight the key function of the striosomes for maintenance of the striatal dopamine tone and contribute to the understanding of their involvement in some neurological disorders such as Huntington's disease.

Usage: Unilateral intrastriatal 2-μl injections of Dermorphin-SAP (17 μg/μl in saline) were performed to induce the selective ablation of MOR-expressing neurons in the striosomal compartment.

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Patch compartment lesions reduce habitual sucrose consumption

Horner KA, Logue JB, Jenrette TA (2017) Patch compartment lesions reduce habitual sucrose consumption. Neuroscience 2017 Abstracts 689.16 / II23. Society for Neuroscience, Washington, DC.

Summary: The striatum mediates habit formation and reward association. The striatum can be divided into the patch and matrix compartment, which are two neurochemically and anatomically distinct regions that may sub-serve different aspects of behavior. For example, the patch compartment may mediate reward-related behaviors, while the matrix compartment may mediate adaptive motor functions. Furthermore, previous studies have shown that enhanced relative activation of the patch versus matrix compartment is associated with inflexible behaviors, such as stereotypy. Habitual behaviors are also inflexible in nature, but whether enhanced activation of the patch compartment contributes to habitual behavior is not known. The goal of the current study was to examine the role of patch compartment neurons in the development of habit formation. We used dermorphin-saporin to specifically ablate neurons of the patch compartment prior to training animals to self-administer sucrose on a random interval schedule of reinforcement, which has been shown to foster habit formation. Our data showed that destruction of the neurons of the patch compartment prevented the reinstatement of sucrose self-administration after sucrose devaluation, indicating that absence of the patch compartment interrupted the development of habitual behavior. Our data also indicate that c-Fos levels were decreased in the dorsolateral striatum (DLS) and sensorimotor cortex (SMC), but increased in dorsomedial striatum (DMS) and prefrontal cortex (PFC) in patch-lesioned animals that did not develop habitual behavior, indicating that diminished habit formation is associated with decreased activation of regions that participate in habitual behavior, and increased in regions associated with goal-directed behaviors. Together, these data indicate that the patch compartment participates in habit formation by altering the flow of information through basal ganglia circuits.

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The rostromedial tegmental nucleus and alcohol addiction

Ye J-H, Fu R, He W (2017) The rostromedial tegmental nucleus and alcohol addiction. Oncotarget 8:18624-18625.. doi: 10.18632/oncotarget.15822

Summary: The authors discuss their work with Dermorphin-SAP (Cat. #IT-12) and their demonstration that damage of RMTg MOR-expressing GABAergic neurons by Dermorphin-SAP increased the intake and preference for alcohol, boosted the expression and slowed down the extinction of alcohol conditioned place preference, and increased locomotion. Microinjection of DS into the RMTg substantially reduced the number of RMTg cells. Importantly, the rats that received DS injection elevated their alcohol intake and preference compared to those that received an injection of Blank-SAP (Cat. #IT-21), which did not cause neuronal damage.

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Ablation of μ opioid receptor-expressing GABA neurons in rostromedial tegmental nucleus increases ethanol consumption and regulates ethanol-related behaviors.

Fu R, Chen X, Zuo W, Li J, Kang S, Zhou L, Siegel A, Bekker A, Ye J (2016) Ablation of μ opioid receptor-expressing GABA neurons in rostromedial tegmental nucleus increases ethanol consumption and regulates ethanol-related behaviors. Neuropharmacology 107:58-67. doi: 10.1016/j.neuropharm.2016.02.027

Summary: In this work the authors investigated cellular mechanisms underlying the aversive effects of alcohol that limit its intake. Previous work has linked synaptic inhibition of dopamine neurons in the ventral tegmental area to this aversion. Rats conditioned to ingest ethanol received bilateral injections totaling 3 pmol of Dermorphin-SAP (Cat. #IT-12) into the rostromedial tegemental nucleus (RTMg). Blank-SAP (Cat. #IT-21) was used as a control. Lesioned animals displayed significantly increased preference for, and intake of ethanol, while showing no change in the desire for sucrose. The results indicate that mu opioid expressing GABAergic neurons in the RTMg are highly involved in the regulation of ethanol consumption.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Blank-SAP (Cat. #IT-21)

Alterations in the rostral ventromedial medulla after the selective ablation of μ-opioid receptor expressing neurons.

Harasawa I, Johansen J, Fields H, Porreca F, Meng I (2016) Alterations in the rostral ventromedial medulla after the selective ablation of μ-opioid receptor expressing neurons. Pain 157:166-173. doi: 10.1097/j.pain.0000000000000344

Summary: The rostral ventromedial medulla (RVM) has both excitatory and inhibitory control over nociceptive neurons in the medullary dorsal horn and spinal cord. Previous work has demonstrated that elimination of mu-opioid receptor-expressing neurons in the RVM reduces stress and injury-induced behavioral hypersensitivity, but the effect of losing these cells on the descending inhibitory system has not been examined. The authors administered 1.2 pmol of Dermorphin-SAP (Cat. #IT-12) to each side of the RVM of rats. Saporin (Cat. #PR-01) was used as a control. Characterization of RVM neurons in lesioned animals showed a reduction in on- and off-cells, but no change in the number of neutral cells. These data indicate that mu-opioid receptor-expressing cells in the RVM are not needed for analgesia produced by activation of RVM neurons.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Selective ablation of the intercalated neurons of the amygdala increased the anxiety-like behavior in the Elevated Plus Maze

Palomares E, Hernandez Perez O, Crespo Ramirez M, Aguilar Roblero R, Fuxe K, Perez de la Mora M (2015) Selective ablation of the intercalated neurons of the amygdala increased the anxiety-like behavior in the Elevated Plus Maze. Neuroscience 2015 Abstracts 694.14/N4. Society for Neuroscience, Chicago IL.

Summary: The intercalated (ITC) islands of the amygdala are clusters of inhibitory neurons that surround the basolateral complex (BLA) and contain a dense population of dopamine D1 and μ-opioid receptors. Lateral ITC (lITC) islands provide feed-forward inhibition to the BLA, whereas medial ITC (mITC) islands form an inhibitory interface between the BLA and central nucleus (CeA), the main output region of the amygdala. Previous studies have shown that ITC neurons play a role in fear extinction. However the functional role of the ITC islands in the un-conditioned anxiety has not been studied. To elucidate the involvement of the ITC islands in the anxiety-like behavior in the Elevated Plus Maze, we bilaterally infused the toxin saporin conjugate with the agonist of the μ-opioid receptors, dermorphine, (SAP-DER; 0.75pmol/0.250µl/lado) in closed proximity to the mITC islands to specifically ablate the neurons of the ITC islands. Behaviorally, SAP-DER injections significantly increased the time that the rats spent in the open arm of the maze as compared with their lesion control group. No effects on locomotion in the open-field test were found. These results suggest that ablate of the ITC neurons results in anxiogenic effects and support ITC neurons play an important role in mediate anxiolytic responses.

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Ablation of the patch compartment reduces cocaine-induced stereotypy

Horner KA, Logan M, Murray RC (2015) Ablation of the patch compartment reduces cocaine-induced stereotypy. Neuroscience 2015 Abstracts 506.23/M12. Society for Neuroscience, Chicago IL.

Summary: Repeated exposure to cocaine (COC) induces stereotypy, which is characterized as inflexible, repetitive behavior. Enhanced relative activation of the patch compartment of the striatum has been shown to positively correlate with the emergence of stereotypy following repeated COC treatment, suggesting that stereotypy may be related to preferential activation of this region. However, the specific contribution of the patch compartment to COC-induced stereotypy following repeated exposure is unknown. To elucidate the involvement of the patch compartment to the development of stereotypy in response to repeated COC exposure, we determined if destruction of this sub-region altered COC-induced behaviors. Animals were bilaterally infused in the striatum with the neurotoxin dermorphin-saporin (DERM-SAP; 17 ng/[[Unsupported Character - Symbol Font ]]l) to ablate the neurons of the patch compartment and allowed to recover for eight days. The animals were given daily injections of COC (25 mg/kg) or saline for one week, followed by a weeklong drug-free period. Animals were then given a challenge dose of COC, placed in activity chambers, observed for 2h and sacrificed. DERM-SAP pretreatment reduced the number of mu-labeled patches in the striatum. DERM-SAP pretreatment significantly reduced the intensity and spatial immobility of COC-induced stereotypy. In support of this observation, increased locomotor activity was seen in DERM-SAP pretreated, COC-treated animals. DERM-SAP pretreatment attenuated COC-induced c-Fos expression in the patch compartment, while enhancing COC-induced c-Fos expression in the matrix compartment. These data indicate that the patch compartment is necessary for repetitive behavior and suggests that alterations in activity in the patch vs matrix compartments may contribute to this phenomenon.

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Striatal patch compartment lesions reduce stereotypy following repeated cocaine administration.

Murray R, Logan M, Horner K (2015) Striatal patch compartment lesions reduce stereotypy following repeated cocaine administration. Brain Res 1618:286-298. doi: 10.1016/j.brainres.2015.06.012

Summary: Stereotypy is defined as abnormally repetitive motor movements accompanied by an inability to initiate normal adaptive responses. Psychostimulants such as cocaine will often produce these movements. It is thought that stereotypy is related to activation of the patch compartment of the striatum. In order to better understand the function of the patch compartment in stereotypy due to repeated exposure to cocaine, the authors administered bilateral injections of Dermorphin-SAP (Cat. #IT-12) into the rostral striatum. Saporin (Cat. #PR-01) was used as a control.

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Endogenous opioid activity in the anterior cingulate cortex is required for relief of pain.

Navratilova E, Xie J, Meske D, Qu C, Morimura K, Okun A, Arakawa N, Ossipov M, Fields H, Porreca F (2015) Endogenous opioid activity in the anterior cingulate cortex is required for relief of pain. J Neurosci 35:7264-7271. doi: 10.1523/JNEUROSCI.3862-14.2015

Summary: There are a number of neuronal circuits involved in the processing of pain, including those that control somatosensory, affective, and cognitive aspects of pain perception. Opioid signaling in the anterior cingulate cortex (ACC) plays a part in pain modulation - this area has also been implicated in the encoding of pain aversiveness. In order to examine the neuronal mechanisms of pain relief and the following reward, the authors of this paper administered 48 ng of Dermorphin-SAP (Cat. #IT-12) into the rostral ACC of rats. Saporin (Cat. #PR-01) was used as a control. The results illuminate the opioid pathway during pain treatment, and the dependence of nucleus accumbens dopaminergic transmission on upstream ACC opioid circuits during pain processing.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Brainstem opioidergic system is involved in early response to experimental SAH.

Cetas J, McFarlane R, Kronfeld K, Smitasin P, Liu J, Raskin J (2015) Brainstem opioidergic system is involved in early response to experimental SAH. Transl Stroke Res 6:140-147. doi: 10.1007/s12975-014-0378-2

Objective: To determine the cause of poor long-term outcomes after Subarachnoid hemorrhage (SAH).

Summary: Failure of the RVM μ-opioid receptor cells to initiate the compensatory CBF response sets the stage for acute and delayed ischemic injury following SAH.

Usage: To lesion medullary neurons expressing the μ-opioid receptor, Dermorphin–SAP was microinjected as a bilateral dose of 0.5 pmol in 500 nL per side (1 pmol in 1 μL total dose and injection volume). Blank–SAP or vehicle was injected in equal volumes and dose as controls.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Blank-SAP (Cat. #IT-21)

Selective ablation of mu opioid receptor expressing gaba neurons in the rostromedial tegmental nucleus promotes ethanol intake

Fu R, Chen X, Zho W, Li J, Ye J-H (2014) Selective ablation of mu opioid receptor expressing gaba neurons in the rostromedial tegmental nucleus promotes ethanol intake. Neuroscience 2014 Abstracts 267.30. Society for Neuroscience, Washington, DC.

Summary: BACKGROUND AND PURPOSE The cellular mechanisms underlying the aversive effect of ethanol that limits its intake are not well understood, although recent evidence has linked aversion with synaptic inhibition of dopamine neurons in the ventral tegmental area. Emerging evidence indicates that the rostromedial tegmental nucleus (RMTg), a newly defined midbrain structure exerts a major GABAergic inhibitory control over midbrain dopamine neurons and encodes aversive stimuli. The RMTg contains mostly GABAergic neurons and with dense μ-opioid receptor (MOR) immunoreactivity. However, the role of RMTg in the regulation of ethanol intake has not been well investigated. EXPERIMENTAL APPROACH We compared voluntary ethanol intake and locomotion in rats with intra-RMTg infusion of dermorphin-saporin or blank saporin. Dermorphin-saporin is a neurotoxin, which could selectively lesion MOR-expressing neurons. We measured ethanol intake in rats given intermittent access to ethanol (20% vol/vol) using a two bottle choice paradigm. We euthanized the rats, dissected their brains and analyzed the glutamic acid decarboxylase67 (GAD67) and MOR protein expression and immunoreactivity immediately following the behavioral test. KEY RESULTS In rats that received intra-RMTg injection of dermorphin-saporin, we observed a robust increase in the intake of and the preference to ethanol, and in the locomotor activity; but a significantly reduced GAD67 and MOR protein expression, as well as a massive loss of neurons with GAD67 and MOR immunoreactivity within the RMTg. We observed no such changes in rats that received injection of blank saporin or saline. Together, These findings indicate that MOR-expressing GABA neurons in the RMTg play a crucial role in the regulation of ethanol consumption, implicating the dysfunction of these neurons likely play a critical role in the pathogenesis of alcoholism, and that these neurons should represent an appropriate target for the development of therapeutic strategies against alcohol use disorders.

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Descending controls modulate inflammatory joint pain and regulate CXC chemokine and iNOS expression in the dorsal horn.

Carr F, Géranton S, Hunt S (2014) Descending controls modulate inflammatory joint pain and regulate CXC chemokine and iNOS expression in the dorsal horn. Mol Pain 10:39. doi: 10.1186/1744-8069-10-39

Summary: Peripheral joint pathology in conditions such as osteoarthritis does not always correlate to the amount of pain experienced, indicating that chronic pain is present. The role of descending facilitation in this form of chronic pain has not been investigated. The authors examined the role of mu opioid receptor-expressing cells in the rostral vental medulla (RVM) in behavioral hypersensitivity seen in joint pain models. Rats received 1.5 pmol of Dermorphin-SAP (Cat. #IT-12) into the RVM. Lesioned animals displayed prolonged attenuation of hypersensitivity, and altered expression of several genes was detected by qPCR, indicating that descending facilitation in the RVM is involved in joint pain behavior.

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Striatal patch compartment lesions alter methamphetamine-induced behavior and immediate early gene expression in the striatum, substantia nigra and frontal cortex

Murray RC, Gilbert YE, Logan AS, Hebbard JC, Horner KA (2014) Striatal patch compartment lesions alter methamphetamine-induced behavior and immediate early gene expression in the striatum, substantia nigra and frontal cortex. Brain Struct Funct 219:1213-1229. doi: 10.1007/s00429-013-0559-x

Summary: In this work, the authors investigated the function of the patch compartment in abnormally repetitive motor actions (stereotypy) in response to a psychostimulant such as methampheta-mine (meth). Rats received bilateral injections of Dermorphin-SAP (Cat. #IT-12; 17 ng) into the rostral striatum. When treated with meth, lesioned animals displayed reduced stereotypy, increased motor activity, and enhanced c-Fos expression.

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Descending facilitation contributes to changes in dorsal horn gene expression in a rat model of inflammatory joint pain.

Carr F, Géranton SM, Hunt SP (2012) Descending facilitation contributes to changes in dorsal horn gene expression in a rat model of inflammatory joint pain. Neuroscience 2012 Abstracts 785.07. Society for Neuroscience, New Orleans, LA.

Summary: Chronic pain is associated with increased excitability and changes in gene expression within the dorsal horn. Descending facilitation from the rostral ventromedial medulla (RVM) is known to contribute to this excitability and behavioural hypersensitivity in a number of pain states. This would suggest that some of the gene changes associated with chronic pain could be driven by descending pathways terminating in the dorsal horn. We have previously demonstrated that ablation of a subset of RVM neurons expressing the mu opioid receptor (MOR) attenuates behavioural hypersensitivity following joint inflammation. The aim of the present study was to combine lesion of the RVM with microarray analysis of the dorsal horn to identify genes regulated by descending facilitation in this pain model. Selective lesion of MOR expressing cells of the RVM was carried out in rats by microinjection of the selective toxin dermorphin-saporin. Non-lesioned controls received vehicle microinjection. 4 weeks after the lesion procedure when depletion of the MOR+ cells was complete, both groups received an injection of 10μl Complete Freund's Adjuvant to the left ankle joint. 7 days later the animals were sacrificed and the ipsilateral quadrant of the dorsal horn of the spinal cord lumbar region (L4-L6) removed. RNA was extracted and microarray analysis carried out using Affymetrix GeneChip Rat Gene 1.0 ST Arrays. Raw data was analysed in R using Bioconductor open source software. Limma testing was applied and a list of genes differentially regulated in animals with prior RVM lesion compared to non-lesioned controls was generated. The majority of differentially regulated genes (73%) were downregulated in the lesioned group. We used the DAVID bioinformatics resource to cluster the genes into groups with similar functional annotations (Huang et al., 2009). This analysis identified 16 gene clusters with significantly enriched functional annotations. Among these enriched functions were ribosomal function and biogenesis, inflammatory response (including the chemokine CXCL10), GPCR signalling (including the serotonin receptor 5HTR1D) and transcriptional regulation (including the transcriptional repressor RCOR2). Following on from identification of functional categories, validation of genes of interest was carried out using RT-qPCR. Our findings suggest that descending facilitation contributes to gene expression changes within the dorsal horn and that this may correlate with behavioural hypersensitivity observed in chronic pain.

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Intercalated nucleus modulates chemosensory processing in medial amygdala.

Biggs LM, Simonton AR, Meredith M (2012) Intercalated nucleus modulates chemosensory processing in medial amygdala. Neuroscience 2012 Abstracts 781.07. Society for Neuroscience, New Orleans, LA.

Summary: The vomeronasal organ is necessary for interpretation by naive rodents (hamsters, mice) of many chemosensory signals. Information is relayed to medial amygdala (Me) via the accessory olfactory bulbs. FRA (Fos related antigen) responses in Me to chemosensory cues suggest this area is important for categorization of cues based on biological relevance to the animal. Anterior Me (MeA) is activated by all chemosensory cues (conspecific and heterospecific). Posterior Me (MeP) activates for conspecific and biologically relevant heterospecific stimuli only. Other heterospecific stimuli suppress MeP, apparently via GABA inhibition, while the adjacent medial-caudal intercalated nucleus (m-ICNc) is activated. Intercalated nuclei (ICNs) are groups of GABAergic cells between amygdala main-divisions. Those adjacent to basolateral and central amygdala (BLA, CeA) are known to mediate BLA, CeA responses via GABA inhibition, modulated by inhibitory DA-D1 receptors on ICN cells. We hypothesize that m-ICNc modulates MeP in a similar manner, as suggested by FRAs data, but this has not yet been tested directly. In hamsters, we show a hyperpolarization of MeP cells and suppression of ongoing spiking in whole-cell slice electrophysiology using in-slice stimulation of m-ICNc. The effect of dopamine and other modulators on this functional relationship is under study with agonists and antagonists. ICN, but not Me, cells carry mu-opioid receptors (MORs). In mice, we use Dermorphin-Saporin, a toxin that selectively destroys MOR+ neurons, to lesion m-ICNc to assess its role in chemosensory responses in MeP. Also in mice, a specific MUP protein in male urine (mMU) facilitates learning of a male’s chemosensory signature by females. We have quantified Me response to high and low molecular weight (HMW, LMW) fractions (LMW lacks protein) and whole mMU using FRAs immediate early gene expression. Preliminary results show no significant difference between HMW, LMW, or whole mMU in Me, however within the BLA (involved in volatile odor learning), there are significant differences in activity between stimuli in females without post-weaning exposure to male urine and no prior exposure to adult male urine.

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Brainstem facilitations and descending serotonergic controls contribute to visceral nociception but not pregabalin analgesia in rats.

Sikandar S, Bannister K, Dickenson AH (2012) Brainstem facilitations and descending serotonergic controls contribute to visceral nociception but not pregabalin analgesia in rats. Neurosci Lett 519(1):31-36. doi: 10.1016/j.neulet.2012.05.009

Summary: Neurons in the rostral ventromedial medulla (RVM) are classified as ON, OFF, or NEUTRAL based on firing patterns in response to noxious somatic stimulation. ON cells express μ-opioid receptors, and are therefore a target for dermorphin-SAP (Cat. #IT-12). The authors injected the RVM of rats with 3 pmol of dermorphin-SAP; Saporin (Cat. #PR-01) was used as a control. Results show the μ-opioid receptor population is not needed for the function of analgesics through the serotonergic system.

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The role of descending facilitation in the initiation and maintenance of mechanical hypersensitivity following inflammation

Carr F, Géranton SM, Hunt SP (2011) The role of descending facilitation in the initiation and maintenance of mechanical hypersensitivity following inflammation. Neuroscience 2011 Abstracts 702.10. Society for Neuroscience, Washington, DC.

Summary: Central sensitisation is the key mechanism involved in the generation of mechanical hypersensitivity associated with tissue injury. Dorsal horn excitability is subject to regulation by descending modulation via the rostral ventromedial medulla (RVM) and enhanced descending facilitation under conditions of persistent nociceptive input contributes to the maintenance of mechanical hypersensitivity in chronic pain states. Depletion of mu-opioid receptor expressing (MOR+) cells of the RVM and depletion of spinal serotonin have been used previously to demonstrate the contribution of descending facilitation to the maintenance of neuropathic pain. Here we have used the same ablation techniques to investigate the contribution of descending pathways to the initiation and maintenance of mechanical hypersensitivity associated with ankle joint inflammation. Male Sprague-Dawley rats (215-220g at the time of injection) received bilateral microinjections of the selective cytotoxin dermorphin-saporin (1.5pM each side). 28 days later the animals received either an injection of 10μl Complete Freund’s Adjuvant (CFA) to the left ankle joint or underwent a sham procedure. Mechanical hypersensitivity of the hindpaw plantar surface was assessed using von Frey hairs from 2 hours up to 8 days post CFA injection. In a separate group of rats (160-180g at the time of injection) depletion of spinal serotonin was out carried out by intrathecal administration of 5,7-dihydroxytrptamine (5,7-DHT). Animals received either 10 μl of 5,7-DHT in saline (6μg/μl) or vehicle control. 6 days later animals received either CFA injection or underwent a sham procedure and mechanical hypersensitivity was assessed as in the dermorphin-saporin experiment. Depletion of the MOR+ cells of the RVM and of spinal serotonin was confirmed using immunohistochemistry. Dermoprhin-saporin pre-treatment resulted in significantly increased paw withdrawal thresholds from 6 hours up to 8 days following CFA injection (p < 0.01, ANOVA with repeated measures). In contrast depletion of spinal serotonin by 5,7-DHT led to a smaller attenuation of mechanical hypersensitivity at 24 hours and 48 hours following inflammation (LSD post hoc test, p < 0.01) but did not result in significantly increased paw withdrawal thresholds at the earlier time points.

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Selective ablation of mu-opioid receptor expressing neurons in the rostral ventromedial medulla attenuates stress-induced mechanical hypersensitivity.

Reynolds J, Bilsky EJ, Meng ID (2011) Selective ablation of mu-opioid receptor expressing neurons in the rostral ventromedial medulla attenuates stress-induced mechanical hypersensitivity. Life Sci 89(9-10):313-9. doi: 10.1016/j.lfs.2011.06.024

Summary: Animals have been shown to develop hyperalgesia in response to chronic stress. Recent data has implicated the rostroventromedial medulla (RVM) in this process. In order to clarify what role mu-opioid receptor expressing neurons in the RVM play in rat, the authors injected 1.8 pmol of dermorphin-SAP (Cat. #IT-12) into the RVM. The rats were then subjected to a model designed to produce hypersensitivity in the hind paw. Stress-induced behavior did not change in the lesioned animals, but mechanical hypersensitivity was reduced.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Neuropeptide receptor co-expression in superficial dorsal horn: Effects of galanin-saporin, neuropeptide y-saporin and dermorphin-saporin

Lemons LL, Chatterjee K, Wiley RG (2010) Neuropeptide receptor co-expression in superficial dorsal horn: Effects of galanin-saporin, neuropeptide y-saporin and dermorphin-saporin. Neuroscience 2010 Abstracts 585.5/XX19. Society for Neuroscience, San Diego, CA.

Summary: We have previously shown that the role of specific neurons in behavioral processes can be fruitfully studied using targeted toxins. Toxins composed of a targeting neuropeptide coupled to the ribosomal-inactivating toxin, saporin, are used to selectively destroy superficial dorsal horn neurons expressing the cognate peptide receptors followed by assessment of changes in pain behavior. In the present study, we sought to compare the anatomic effects of three closely related targeted toxins, each with different nocifensive behavioral effects. Rats were given single lumbar intrathecal injections of either galanin-saporin (Gal-sap), neuropeptide Y-saporin (NPY-sap), or dermorphin-saporin (Derm-sap). Lumbar spinal cord sections from each rat were stained for each of the three receptors, GalR-1, Y1R and MOR (mu opiate) using standard immunoperoxidase technique. Each toxin produced a significant decrease in staining for its cognate receptor. Gal-sap animals showed no change in either MOR or Y1R staining. NPY-sap rats showed decreased staining for both GalR1 and MOR, and Derm-sap rats were assessed for changes in expression of GalR1 and Y1R. These findings suggest overlaps between the populations of neurons that express the GalR1, Y1R, and MOR. Specifically, Y1R-expressing neurons also express GalR1 and MOR, probably by separate subpopulations of Y1R neurons. The results also suggest either that Gal-sap only kills neurons that do not express either of the other two receptors, or some of the observed loss of receptors after NPY-sap is due to secondary (transsynaptic) effects. Double- and triple-label fluorescent immunohistochemistry will be used to directly visualize receptor co-expression patterns and targeted toxin effects. These results will be valuable in interpreting the unique nocifensive behavioral effects of each of these targeted toxins.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Galanin-SAP (Cat. #IT-34), NPY-SAP (Cat. #IT-28)

Spinal µ-opiate receptor (MOR)-expressing dorsal horn neurons: Role in modulating pain and opiate analgesia.

Kline IV RH, Wiley RG (2009) Spinal µ-opiate receptor (MOR)-expressing dorsal horn neurons: Role in modulating pain and opiate analgesia. Neuroscience 2009 Abstracts 560.13/CC72. Society for Neuroscience, Chicago, IL.

Summary: Selective destruction of MOR-expressing interneurons in lamina II of the dorsal horn of the spinal cord increases reflex nocifensive responses to formalin and decreases the anti-nociceptive effects of morphine on the hotplate and in the formalin test. The interpretation of these studies is limited because reflex-based assays may not accurately reflect the cerebral component of nociception. Therefore, we sought to determine the effects of selectively destroying MOR-expressing dorsal horn neurons on baseline operant responses to aversive thermal and mechanical stimuli in a shuttle box task and effects of systemic morphine and naloxone in the same task. The preference apparatus consisted of a 15 X 15 X 30 cm smoked Plexiglas vented chamber placed upon two adjoining temperature-controlled smooth aluminum floor plates (thermal preference task) or one smooth temperature-controlled floor plate adjoined to a room temperature surface covered with 40 grit sandpaper (mechanical preference task). For both preference tasks, response functions were obtained by pairing a 44°C plate or the sandpaper surface with either 11°, 16°, 25°, 38° or 46°C. Rats were intrathecally injected over the lumbar cord with either 625ng of derm-sap (n=7) or blank-sap (n=6) followed by daily thermal or mechanical preference testing on a randomized schedule. Derm-sap treated rats showed enhanced avoidance of aversive thermal stimuli and the aversive mechanical stimulus. Morphine and naloxone significantly altered responses of control rats (blank-sap), but not derm-sap rats, in both thermal and mechanical preference tasks. We interpret these results as showing that the derm-sap lesion produces hyperalgesia/allodynia, impairs the anti-nociceptive and analgesic effects of morphine and therefore indicating that postsynaptic dorsal horn MOR-expressing neurons play a key role in modulating nociception, pain and opiate analgesia. Dysfunction of these neurons may also play a role in pathological pain states.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Partial ablation of mu-opioid receptor rich striosomes produces deficits on a motor-skill learning task.

Lawhorn C, Smith DM, Brown LL (2009) Partial ablation of mu-opioid receptor rich striosomes produces deficits on a motor-skill learning task. Neuroscience 163(1):109-119. doi: 10.1016/j.neuroscience.2009.05.021

Summary: The functional role of basal ganglia striosomes is not well understood. In order to examine these cells in the context of motor behavior the authors injected 8.5 ng of dermorphin-SAP (Cat. #IT-12) into several areas of the striatum of mice (saporin, Cat. #PR-01, was used as a control). The animals were then evaluated in complex motor tasks involving the use of striatal circuitry. Animals receiving dermorphin-SAP showed deficits in specific motor tasks corresponding to the extent of the lesion.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Evaluation of side effects through selective ablation of the mu opioid receptor expressing descending nociceptive facilitatory neurons in the rostral ventromedial medulla with dermorphin-saporin.

Cao F, Chen SS, Yan XF, Xiao XP, Liu XJ, Yang SB, Xu AJ, Gao F, Yang H, Chen ZJ, Tian YK (2009) Evaluation of side effects through selective ablation of the mu opioid receptor expressing descending nociceptive facilitatory neurons in the rostral ventromedial medulla with dermorphin-saporin. Neurotoxicology 30(6):1096-1106. doi: 10.1016/j.neuro.2009.06.004

Summary: Selective ablation of rostral ventromedial (RVM) neurons expressing mu opioid receptors has been suggested as a treatment for pathological pain. This work investigated the side effects of a 0.5 µg injection of dermorphin-SAP (Cat. #IT-12) into the RVM. Saporin (Cat. #PR-01) was used as a control. Lesioned animals experienced a temporary increase in heart rate and systolic blood pressure, and mild microglial responses, but even these soon returned to normal. The data suggest this system has potential as a target for pain therapeutics.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Recent progress in research on ribosome inactivating proteins.

Ng TB, Wong JH, Wang H (2009) Recent progress in research on ribosome inactivating proteins. Curr Protein Pept Sci 1323:94-108. doi: 10.2174/138920310790274662

Summary: This review discusses recent literature on ribosome inactivating proteins including the use of saporin-based conjugates in neuroscience and cancer research. Brief descriptions of research done using 192-IgG-SAP (Cat. #IT-01), OX7-SAP (Cat. #IT-02), dermorphin-SAP (Cat. #IT-12), anti-SERT-SAP (Cat. #IT-23), SSP-SAP (Cat. #IT-11), anti-DBH-SAP (Cat. #IT-03), CTB-SAP (Cat. #IT-14), and other conjugates are provided along with basic information about ribosome inactivating proteins.

Related Products: 192-IgG-SAP (Cat. #IT-01), OX7-SAP (Cat. #IT-02), Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Anti-SERT-SAP (Cat. #IT-23), SSP-SAP (Cat. #IT-11), Anti-DBH-SAP (Cat. #IT-03), CTB-SAP (Cat. #IT-14)

Sex differences in micro-opioid receptor expression in the rat midbrain periaqueductal gray are essential for eliciting sex differences in morphine analgesia.

Loyd DR, Wang X, Murphy AZ (2008) Sex differences in micro-opioid receptor expression in the rat midbrain periaqueductal gray are essential for eliciting sex differences in morphine analgesia. J Neurosci 28:14007-14017. doi: 10.1523/JNEUROSCI.4123-08.2008

Summary: The authors test whether the periaqueductal gray (PAG), that contains a dense population of µ-opioid receptor (MOR)-expressing neurons, is sexually dimorphic. Rats were injected with 3 pmol of Dermorphin-SAP (Cat. #IT-12) into the PAG. Blank-SAP (Cat. #IT-21) was used as a control. Both behavioral and immunohistochemical evidence suggest that differential expression of MOR-expressing neurons in the PAG between male and female rats accounts for the difference in response to morphine.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Blank-SAP (Cat. #IT-21)

Neuropathic pain is maintained by brainstem neurons co-expressing opioid and cholecystokinin receptors.

Zhang W, Gardell S, Zhang D, Xie JY, Agnes RS, Badghisi H, Hruby VJ, Rance N, Ossipov MH, Vanderah TW, Porreca F, Lai J (2009) Neuropathic pain is maintained by brainstem neurons co-expressing opioid and cholecystokinin receptors. Brain 132:778-787. doi: 10.1093/brain/awn330

Summary: It has been hypothesized that a subset of rostral ventromedial medulla (RVM) neurons co-expressing the cholecystokinin type 2 receptor and the mu-opioid receptor are responsible for the maintenance of neuropathic pain. Rats were treated with 50-ng bilateral RVM injections of Dermorphin-SAP (Cat. #IT-12), CCK-SAP (Cat. #IT-31), or saporin (Cat. #PR-01) as a control. Lesion of the RVM neurons prevented hyperalgesia in response to CCK treatment, and shortened abnormal pain states caused by sciatic nerve injury.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), CCK-SAP (Cat. #IT-31), Saporin (Cat. #PR-01)

Analysis of inhibitory phase of formalin test: Effects of specific neural lesions

Wiley RG, Moore SA, Kline IV RH (2008) Analysis of inhibitory phase of formalin test: Effects of specific neural lesions. Neuroscience 2008 Abstracts 772.4/MM19. Society for Neuroscience, Washington, DC.

Summary: The formalin test has been widely used as a model of persistent pain. The 90 mins of formalin-induced nocifensive responding can be divided into two phases (phase 1, first ~10 mins; phase 2, last ~60 mins) separated by a period of reduced responding (interphase, IP), that has received relatively little attention. Behavioral inhibition during the IP of the formalin test has been associated with electrophysiological evidence of inhibition of dorsal horn nociceptive neurons (Henry et al, Pain, 82:57, 1999), probably due, at least in part, to local spinal mechanisms. Behavioral inhibition during IP has been shown to be enhanced by morphine and suppressed by naloxone. In the present study, we sought to determine the effect of selective depletion of specific dorsal horn interneurons known to be involved in nociception, i.e. neurons expressing NPY1R, GalR1 or MOR, or selective destruction of cerebral noradrenergic neurons or spinal cord projecting 5-HT neurons on formalin-induced nociceptive behavior, with particular attention to IP. Type-selective lesions were produced by lumbar intrathecal injection of NPY-saporin, galanin-saporin or dermorphin-saporin, respectively. Cerebral noradrenergic neurons and spinally projecting 5-HT neurons were destroyed using the immunotoxins, antiDBH-saporin (intracerebroventricular) or antiSERT-saporin (lumbar intrathecal), respectively. Partial loss of dorsal horn interneurons expressing NPY1R or GalR1 decreased nocifensive responding during IP and phase 2 of the formalin test, while partial loss of MOR-expressing dorsal horn interneurons increased nocifensive responding during IP and during phase 2. Both antiDBH-sap and antiSERT-sap decreased responding during IP, without effects on either phase 1 or 2. These results suggest that the apparent anti-nociception during IP and phase 2 produced by loss of NPY1R- and GalR1-expressing dorsal horn neurons is due to increased inhibition over excitation/facilitation of nociceptive projection neurons, whereas depletion of MOR-expressing interneurons produces the opposite effect. The apparent enhanced nociception during IP, but not phase I and II, produced by antiDBH-sap and antiSERT-sap suggests that these neural systems serve to enhance the excitability of nociceptive projection neurons during the formalin IP. Electrophysiologic and pharmacologic studies of formalin IP in selectively lesioned animals combined with the above behavioral findings may reveal new insights into endogenous modulation of nocifensive motor responses and/or nociception.

Related Products: NPY-SAP (Cat. #IT-28), Anti-SERT-SAP (Cat. #IT-23), Galanin-SAP (Cat. #IT-34), Anti-DBH-SAP (Cat. #IT-03), Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Lumbosacral-bulbo-spinal loop relayed by RVM on-cells regulates visceral nociception and modulates inhibitory effects of pregabalin and ondansetron

Sikandar S, Dickenson AH (2008) Lumbosacral-bulbo-spinal loop relayed by RVM on-cells regulates visceral nociception and modulates inhibitory effects of pregabalin and ondansetron. Neuroscience 2008 Abstracts 269.4/GG19. Society for Neuroscience, Washington, DC.

Summary: Descending controls from brainstem nuclei including the rostral ventromedial medulla (RVM) have been shown to play an important role in visceral pain, and compounds modulating serotonergic receptor activity and compounds targeting the α2δ subunit of voltage-gated calcium channels have demonstrated clinical efficacy in providing symptomatic relief in patients with visceral hyperalgesia. We investigated the role of RVM on-cells and a serotonergic lumbosacral-bulbo-spinal loop in visceral hyperalgesia and examined the antihyperalgesic effects of ondansetron and pregabalin in modifying visceral pain responses to colorectal distension (CRD) in rats. An in vivo model of visceral pain was established involving CRD and a reliable EMG recording protocol for measuring activity in the external oblique muscle following CRD for quantifying evoked visceromotor responses (VMR) in Sprague-Dawley rats. Changes in VMR evoked by CRD in a range of 10-80 mmHg were recorded following administration of ondansetron (50 μg/kg i.t.) and pregabalin (30 mg/kg s.c.) in naïve rats and rats pretreated with 0.25% intracolonic mustard oil (MO) to induce colonic hyperalgesia. Moreover, RVM on-cells were selectively ablated with injection of the neurotoxin saporin conjugated to the μ-receptor agonist dermorphin (DermSAP) using stereotaxic techniques. Twenty-eight days post-injection, the VMR were compared between naïve, SAP and Derm-SAP rats in control conditions and following intracolonic MO. CRD produced graded VMR responses that were facilitated by intracolonic MO. Both ondansetron and pregabalin were shown to effectively reduce evoked VMR to CRD in naïve rats and MO pretreated rats by antagonizing spinal 5-HT3 receptors and by binding to the α2δ subunit of voltage-gated calcium channels, respectively. Moreover, DermSAP pretreatment was shown to reduce overall evoked VMR, and the antihyperalgesic efficacies of ondansetron and pregabalin were also shown to be modified by the loss of on-cells in DermSAP rats. Furthermore, we verified immunohistochemically RVM on-cell ablation in DermSAP rats and quantified RVM 5-HT cell intensity between naïve, SAP and DermSAP rats. This study illustrates the role of 5-HT3-mediated descending facilitatory controls in visceral pain as well as providing evidence for the antihyperalgesic efficacy of the second generation α2δ ligand pregabalin in the CRD model. Moreover, evidence is provided for a facilitatory serotonergic lumbosacral-bulbo-spinal loop relayed by RVM on-cells that is evoked by CRD and modulates efficacies of pregabalin and ondansetron.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Descending facilitation from the brainstem determines behavioural and neuronal hypersensitivity following nerve injury and efficacy of pregabalin.

Bee LA, Dickenson AH (2008) Descending facilitation from the brainstem determines behavioural and neuronal hypersensitivity following nerve injury and efficacy of pregabalin. Pain 140:209-223. doi: 10.1016/j.pain.2008.08.008

Summary: Rostral ventromedial medulla (RVM) facilitatory On cells are thought to be involved in the mechanisms that control chronic pain. Dermorphin-SAP (Cat. #IT-12, 3 pmol injected into the RVM of rats) was used to examine how mu-opioid receptor expressing facilitatory cells fit into this circuit. Saporin (Cat. #PR-01) was used as a control. The results show that activity in the RVM may influence the outcome of nerve injury.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Amygdala intercalated neurons are required for expression of fear extinction.

Likhtik E, Popa D, Apergis-Schoute J, Fidacaro GA, Pare D (2008) Amygdala intercalated neurons are required for expression of fear extinction. Nature 454(7204):642-645. doi: 10.1038/nature07167

Summary: Scientists have been using fear learning in animals to study human anxiety disorders. In order to investigate the contribution of amygdala plasticity to fear learning, rats received 0.25-µl bilateral infusions of 3-µM dermorphin-SAP (Cat. #IT-12) into the amygdala. Blank-SAP (Cat. #IT-21) was used as a control. Lesioned rats displayed extinction expression deficits, indicating that the eliminated intercalated amygdala neurons play a large role in the extinction process.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Blank-SAP (Cat. #IT-21)

Read the featured article in Targeting Trends.

Spinal mu-opioid receptor-expressing dorsal horn neurons: role in nociception and morphine antinociception.

Kline IV RH, Wiley RG (2008) Spinal mu-opioid receptor-expressing dorsal horn neurons: role in nociception and morphine antinociception. J Neurosci 28:904-913. doi: 10.1523/JNEUROSCI.4452-07.2008

Summary: The authors used Dermorphin-SAP (Cat. #IT-12) to investigate the function of spinal cord mu-opioid receptor (MOR)-expressing dorsal horn neurons in nociception and morphine analgesia. Rats were treated with 500 ng intrathecal injections of Dermorphin-SAP; 500 ng of Blank-SAP (Cat. #IT-21), and up to 1 µg of Saporin (Cat. #PR-01) were used as controls. The data indicate that MOR-expressing dorsal horn neurons are necessary for morphine action and play a role in nocifensive responses to persistent pain in the formalin test.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Blank-SAP (Cat. #IT-21), Saporin (Cat. #PR-01)

Selective ablation of GABA neurons in the ventral tegmental area increases spontaneous locomotor activity.

Shank EJ, Seitz PK, Bubar MJ, Stutz SJ, Cunningham KA (2007) Selective ablation of GABA neurons in the ventral tegmental area increases spontaneous locomotor activity. Behav Neurosci 121:1224-1233. doi: 10.1037/0735-7044.121.6.1224

Summary: To further examine the importance of the ventral tegmental area (VTA) g-aminobutyric acid (GABA) neurons in behavioral function, the authors lesioned the VTA of rats with dermorphin-saporin (Cat. #IT-12). Lesioned animals received 1 or 2 pmol/200 nl bilateral injections of conjugate; blank-SAP (Cat. #IT-21) was used as a control. Rats treated with dermorphin-SAP displayed significantly elevated motility as compared to control animals. Rats receiving 1 pmol of dermorphin-SAP returned to normal motility 14 days after treatment, but rats receiving 2 pmol maintained the increased motility through day 14.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Blank-SAP (Cat. #IT-21)

Lesioning mu opioid receptor-containing neurons in the ventrolateral periaqueductal gray attenuates morphine analgesia in male but not female rats

Loyd DR, Murphy AZ (2007) Lesioning mu opioid receptor-containing neurons in the ventrolateral periaqueductal gray attenuates morphine analgesia in male but not female rats. Neuroscience 2007 Abstracts 921.4/NN15. Society for Neuroscience, San Diego, CA.

Summary: Chronic pain will affect four out of five persons at some point across the lifespan. While the opioid-based narcotic morphine is the most prevalent treatment for chronic pain in clinical settings, it is becoming increasingly clear that morphine produces a significantly greater degree of analgesia in males compared to females. In both somatic and visceral pain models, the ED50 for morphine is generally two-fold higher for females than for males. The midbrain periaqueductal gray (PAG) and its descending projections to the rostral ventromedial medulla (RVM) is the primary circuit for opioid-based analgesia. We have recently shown that the PAG-RVM pathway is sexually dimorphic both in its anatomical organization and in its activation during persistent pain. Interestingly, while female rats have a greater number of PAG neurons that project to the RVM, inflammatory pain activates these cells to a greater degree in males. Additionally, systemic morphine inhibits the pain-induced activation of PAG neurons in males, but not females. Sex differences in neuronal activity during pain and morphine analgesia are prominent in the ventrolateral PAG, a region containing a large population of mu opioid receptor-containing neurons. We have recently shown that females have significantly lower levels of mu opioid receptors (MOR) in this region, however it is not known whether sex differences in MOR expression contribute to our observed sex differences in morphine analgesia. To test the role of ventrolateral PAG MOR in morphine analgesia, the cytotoxin saporin conjugated to the MOR agonist dermorphin (Der-Sap) was injected into the ventrolateral PAG to site-specifically lesion MOR-containing neurons. Twenty-eight days later, rats received an intraplantar injection of CFA to induce persistent pain and twenty-four hours later morphine was administered systemically using a cumulative dosing paradigm (1.8 -18mg/kg). Lesions of PAG MOR-containing neurons resulted in a two-fold rightward shift in morphine ED50 values in male rats compared to controls. Interestingly, in females no difference was noted in morphine ED50 for Der-Sap treated females versus controls suggesting that the PAG is not a critical site for morphine analgesia in females. Der-Sap treatment had no significant impact on baseline paw withdrawal latencies or CFA-induced hyperalgesia. These results indicate that the PAG is a primary locus for systemic morphine analgesia in males only and suggests the necessity for the development of sex-specific treatments for persistent pain in females.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Role of spinal cord µ-opioid receptor expressing dorsal horn neurons in morphine analgesia

Kline IV RH, Wiley RG (2006) Role of spinal cord µ-opioid receptor expressing dorsal horn neurons in morphine analgesia. Neuroscience 2006 Abstracts 643.19. Society for Neuroscience, Atlanta, GA.

Summary: The role of spinal cord μ-opioid receptor expressing dorsal horn neurons in morphine analgesia is not clearly understood. Using lumbar intrathecal (i.t.) injections of the targeted toxin dermorphin-saporin to selectively destroy these cells, we sought to determine the effect of this lesion on the antinociceptive activity of systemic and i.t. morphine on the hotplate test. We examined the antinociceptive effects of morphine across a range of stimulus intensities (44, 47 & 52oC) in order to assess responses mediated by C or Aδ thermal nociceptors. Experiment 1 (systemic morphine): Sixteen Sprague Dawley male rats were injected with 500ng dermorphin-saporin i.t. or PBS and hotplate testing resumed one week after injections. Baseline hotplate responses were monitored for three weeks after which systemic morphine dose response curves (0, 2.5, 5, &10 mg/kg s.c.) were performed. Experiment 2 (spinal intrathecal morphine): Twelve Long Evans female rats were surgically implanted with indwelling lumbar i.t. catheters (8.5cm), underwent baseline hotplate testing for 7 days, had i.t. morphine dose response curves (0, 0.01, 0.1, & 1 μg) performed at 44 & 52oC seven days before and eight days after dermorphin-saporin injections. The dependent measures for the hotplate test were: 1) latencies to the first lick or guard response (all temperatures) and 2) the cumulative durations and amounts of licking and guarding events (44 and 47oC). Loss of lamina II MOR-expressing dorsal horn neurons after dermorphin-saporin was confirmed in spinal cord sections from each rat stained for MOR1 and MOR1C using standard immunoperoxidase techniques on adjacent 40 μm sections from the L4 spinal segment. Baseline responses to noxious heat did not decrease after i.t. dermorphin-saporin. The antinociceptive activity of systemic morphine was attenuated in dermorphin-saporin treated rats at 44 & 47oC; this effect was least striking on the 52oC hotplate and greatest on the 44oC hotplate. The dermorphin-saporin-induced lesion reduced the antinociceptive effects of intrathecal morphine more than systemic morphine. Based on the above findings are others not included here, we conclude that dorsal horn MOR expressing neurons are necessary for morphine to exert its maximum antinociceptive and analgesic effects.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

A selective lesioning method to probe the role of intercalated (ITC) amygdala neurons in the extinction of classically conditioned fear responses

Likhtik E, Apergis-Schoute J, Pare D (2006) A selective lesioning method to probe the role of intercalated (ITC) amygdala neurons in the extinction of classically conditioned fear responses. Neuroscience 2006 Abstracts 370.19. Society for Neuroscience, Atlanta, GA.

Summary: The acquisition of conditioned fear responses (CRs) is thought to involve the potentiation of synapses conveying information about the conditioned stimulus (CS) to the basolateral (BLA) amygdala. Expression of CRs would depend on transfer of potentiated CS inputs by the BLA to the central amygdala (CE). In contrast, the mechanisms of extinction remain controversial. It was proposed that ITC neurons, which receive BLA inputs and generate feedforward inhibition in CE, are in a key position to mediate extinction. In this view, potentiation of BLA inputs to ITC cells during extinction training, would dampen the impact of CS-related BLA activity on CE neurons, inhibiting CRs. However, this idea is difficult to test because ITC cells occur in small, lateromedially dispersed clusters, making conventional lesioning methods inadequate. The present study aimed to find an effective way of eliminating ITC cells, taking advantage of the fact that, compared to the rest of the amygdala, they exhibit strong immunoreactivity for mu opioid receptors (muORs). First, we performed electron microscopic observations to determine whether muORs are expressed by ITC cells vs. afferents to ITC cells. This test revealed that muORs are concentrated in the postsynaptic membrane density of asymmetric synapses found on ITC cells. Next, we tested whether it is possible to obtain selective ITC lesions by injecting the toxin saporin conjugated to the mu opioid agonist dermorphin (DER-SAP) in the proximity of ITC cells. Thus, rats received intra-amygdaloid pressure injections of DER-SAP in one hemisphere and of vehicle on the contralateral side. Seven days later, the animals were perfused and the tissue processed to reveal muOR. DER-SAP injections produced a marked reduction in muOR immunoreactivity at the BLA-CE border, where ITC cells are usually located. Thus, selective lesioning of ITC cells can be achieved using this method. We are currently testing the impact of such ITC lesions on extinction learning.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Descending facilitation from the rostral ventromedial medulla maintains nerve injury-induced central sensitization.

Vera-Portocarrero LP, Zhang ET, Ossipov MH, Xie JY, King T, Lai J, Porreca F (2006) Descending facilitation from the rostral ventromedial medulla maintains nerve injury-induced central sensitization. Neuroscience 140(4):1311-1320. doi: 10.1016/j.neuroscience.2006.03.016

Summary: Rats were treated with 1.5 pmol of dermorphin-SAP (Cat. #IT-12) or saporin (Cat. #PR-01) into each side of the rostral ventromedila medulla, followed by spinal nerve ligation. The data indicate that mu opioid-expresing neurons are necessary to maintain nerve injury-induced central sensitization.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Saporin (Cat. #PR-01)

Descending facilitation from the rostral ventromedial medulla maintains visceral pain in rats with experimental pancreatitis.

Vera-Portocarrero LP, Yie JX, Kowal J, Ossipov MH, King T, Porreca F (2006) Descending facilitation from the rostral ventromedial medulla maintains visceral pain in rats with experimental pancreatitis. Gastroenterology 130(7):2155-2164. doi: 10.1053/j.gastro.2006.03.025

Summary: Here the authors investigated the role of ascending or descending pathways in the mediation of pain caused by pancreatitis. Rats received 1.5 pmol injections of dermorphin-SAP (Cat. #IT-12) into each side of the rostral ventromedial medulla. Abdominal hypersensitivity was tested using von Frey filaments. Although the ablation of mu-opioid receptor-expressing neurons by dermorphin-SAP did not prevent the initial expression of pancreatitis pain, maintenance of this pain was absent. The data link maintenance of pancreatitis pain to descending pathways.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Increased formalin behavior after selective destruction of μ opiate receptor-expressing dorsal horn neurons: impaired descending analgesic control?

Datta S, Kline IV RH, Wiley RG (2005) Increased formalin behavior after selective destruction of μ opiate receptor-expressing dorsal horn neurons: impaired descending analgesic control?. Neuroscience 2005 Abstracts 623.15. Society for Neuroscience, Washington, DC.

Summary: Spinal intrathecal injection of dermorphin-saporin (derm-sap) selectively destroys dorsal horn neurons expressing the mu-opiate receptor (MOR). In the present study, we sought to determine the effect of derm-sap (500 ng, i.t.) on responses to intraplantar formalin injection (25 ul of 5%). After formalin injection, rats were immediately placed into a clear observation chamber with a video camera beneath the floor. Rats were videotaped for 90 minutes and their behavior scored offline for one minute out of every 5 minutes. 120 minutes after formalin injection rats were anesthetized with pentobarbital and perfused with formalin. Spinal cord sections were stained for MOR and cholecystokinin (CCK) using standard immunoperoxidase techniques on adjacent 40 um sections from L4 spinal segment. Coded sections were used to assess MOR staining intensity by quantitative densitometry. Derm-sap treated rats showed no separation between phase I and II and spent more time than vehicle controls licking/guarding/biting the injected hindpaw during both phase I and II. Derm-sap significantly decreased dorsal horn MOR. Staining for CCK showed time dependant changes after derm-sap which was not present in PBS controls. These same derm-sap treated rats performed normally on hotplate at 44, 47 and 52 C and had normal analgesic responses to systemic morphine on 44, 47 and 52 C hotplates. We interpret these data to indicate that loss of the dorsal horn MOR-expressing neurons reduces the effect of descending analgesic mechanisms. Supported by NIH R21-DA14380 and Department of Veterans Affairs.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Facilitatory influences from the rostral ventromedial medulla (RVM) are required for pancreatic nociception

Vera-Portocarrero LP, Xie Y, King T, Lai J, Porreca F (2005) Facilitatory influences from the rostral ventromedial medulla (RVM) are required for pancreatic nociception. Neuroscience 2005 Abstracts 623.18. Society for Neuroscience, Washington, DC.

Summary: Pain is a frequent complaint of patients with pancreatitis or pancreatic cancer. An animal model of pancreatitis induced by dibutyltin dichloride (DBTC) is characterized by abdominal hypersensitivity to mechanical stimuli that appears by day 3 after induction of pancreatitis and persists for at least 10 days. We have used this model to evaluate the role of descending pain modulatory pathways from the RVM in the processing of visceral pain. Pancreatitis was induced in rats by a single tail vein injection of DBTC. Animals were monitored for mechanical sensitivity of the abdominal area as an index of pancreatic nociception using von Frey hairs applied to the surface of the abdomen and recording the frequency of withdrawals from stimulation. Six days after DBTC injection, when mechanical hypersensitivity was fully developed, lidocaine, or saline, was microinjected into the RVM. Lidocaine, but not saline, given into the RVM produced a time-related reversal of mechanical hypersensitivity which peaked by 20 min after injection in animals with pancreatitis. RVM lidocaine had no effect on rats without pancreatitis. A second group of rats received a single microinjection of the cytotoxin dermorphin-saporin into the RVM in order to ablate mu opioid receptor expressing cells that have been proposed to drive descending pain facilitation. 28 days later, the rats received DBTC and their response to mechanical stimulation was monitored daily. These rats showed mechanical hypersensitivity on day 3 after DBTC, but the sensory threshold reverted to normal level by day 6, while rats that had been pretreated with dermorphin, saporin, or water exhibited persistent mechanical hypersensitivity after DBTC out to day 10. These data suggest that a blockade of the descending input from the RVM by lidocaine is sufficient to block the pancreatitis-induced visceral pain, and that the mu opioid receptor expressing cells in the RVM are critical for the persistent pain state.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Pain facilitatory cells in the rostral ventromedial medulla coexpress opioid-μ receptors and cholecystokinin type 2 receptors

Zhang W, Gardell SE, Xie Y, Luo M, Rance NE, Vanderah TW, Porreca F, Lai J (2005) Pain facilitatory cells in the rostral ventromedial medulla coexpress opioid-μ receptors and cholecystokinin type 2 receptors. Neuroscience 2005 Abstracts 394.17. Society for Neuroscience, Washington, DC.

Summary: Pain transmission can be modulated by descending input to the spinal dorsal horn from the rostral ventromedial medulla (RVM). RVM neurons that facilitate nociception are termed “ON-cells”, which are inhibited by mu-opioids, suggesting that they express opioid mu receptors (MOR). Focal application of cholecystokinin (CCK8(s)) into the RVM elicits acute thermal and tactile hypersensitivity and induces ON-cell activity. In situ hybridization using riboprobes for either rat MOR or rat cholecystokinin type-2 receptor (CCK-2) confirms the expression of these receptors in the RVM. Pretreatment with a toxin conjugate, CCK8(s)-saporin results in a significant loss of CCK-2 positive cells in the RVM, concomitant with a blockade of CCK8(s) induced hyperalgesia. The pretreatment also significantly reduces the number of neurons labeled for MOR in the RVM, suggesting that MOR and CCK-2 may be co-localized in some RVM cells. Consistent with these data, similar pretreatment with the toxin conjugate, dermorphin-saporin, which selectively targets MOR expressing neurons, significantly reduces the number of MOR labeled cells in the RVM, blocks RVM CCK8(s) induced hyperalgesia and reduces the number of CCK-2 positive cells in the RVM. In situ hybridization using 35S-labeled CCK-2 riboprobes and Digoxigenin-labeled MOR riboprobes shows that over 80% of labeled RVM neurons co-express both MOR and CCK-2, ~15% express only CCK-2, and very few cells express only MOR. These findings represent the first direct demonstration of the phenotype of pain facilitatory neurons in the RVM. Together with previous studies showing that RVM CCK-2 antagonists reverse nerve injury-induced pain, this phenotype provides strong support for the view that endogenous CCK is a critical mediator of the descending pain facilitation, particularly in the maintenance of experimental neuropathic pain. Support Contributed By: NIDCR R01 DE016458

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12), CCK-SAP (Cat. #IT-31)

ATS Poster of the Year Winner

Molecular neurosurgery with targeted toxins

Wiley RG, Lappi DA (2005) Molecular neurosurgery with targeted toxins. Humana Press, Totowa, New Jersey.

Summary: The idea behind the book was to provide a road map for the users of Molecular Neurosurgery to see how experienced scientists used these exceptional reagents in their work. Experiments with several targeted toxins are described, and readers can get an idea either specifically about a targeted toxin that they’re using, or about how a type of molecule is used and at what dosage, in a paradigm similar to theirs.

Related Products: 192-IgG-SAP (Cat. #IT-01), ME20.4-SAP (Cat. #IT-15), Anti-DBH-SAP (Cat. #IT-03), SSP-SAP (Cat. #IT-11), Dermorphin-SAP / MOR-SAP (Cat. #IT-12), IB4-SAP (Cat. #IT-10), CTB-SAP (Cat. #IT-14)

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Mu opioid receptor-containing neurons mediate electroacupuncture-produced anti-hyperalgesia in rats with hind paw inflammation.

Zhang RX, Wang L, Liu B, Qiao JT, Ren K, Berman BM, Lao L (2005) Mu opioid receptor-containing neurons mediate electroacupuncture-produced anti-hyperalgesia in rats with hind paw inflammation. Brain Res 1048(1-2):235-240. doi: 10.1016/j.brainres.2005.05.008

Summary: Electroacupuncture has been shown to significantly reduce inflammatory hyperalgesia. To examine whether this effect is modulated by spinal mu opioid receptors, the authors injected 400 ng of dermorphin-SAP (Cat. #IT-12) into the subarachnoid space at the level of the lumbar spinal cord of rats. The anti-hyperalgesic effect of electroacupuncture was blocked by dermorphin-SAP administration, indicating that mu opioid receptor-containing neurons are involved in this pathway.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Mu-opioid receptor-expressing neurons in the nucleus reticularis gigantocellularis contribute to descending facilitation during the development of inflammatory pain

Wei F, Zou S, Robbins MT, Ren K, Dubner R (2004) Mu-opioid receptor-expressing neurons in the nucleus reticularis gigantocellularis contribute to descending facilitation during the development of inflammatory pain. Neuroscience 2004 Abstracts 297.3. Society for Neuroscience, San Diego, CA.

Summary: We have previously shown that nucleus reticularis giangocellularis (NGC) is involved in descending facilitation of inflammatory hyperalgesia. The cellular mechanisms of descending facilitation from the NGC are unknown. The targeted destruction of the mu-opioid receptor-containing neurons in the rostral ventromedial medulla (RVM) by a dermorphin-saporin conjugate prevents nerve injury-induced hyperalgesia in rats (Porreca et al., J. Neurosci. 21:5281, 2001). We examined the effects of selective deletion of the mu-opioid receptor-expressing neurons in the sub-regions of RVM on nocifensive behaviors in rats. After microinjection of dermorphin-saporin conjugate (1.5 pmol/500 nl) into the RVM, there were no changes in baseline thermal and mechanical sensitivity to noxious stimuli. However, the injection of dermorphin-saporin conjugate into bilateral NGC (n=7) significantly attenuated the thermal hyperalgesia and mechanical allodynia at 30 min to 1 d after hindpaw inflammation produced by injection of complete Freund’s adjuvant, compared to sham (blank-saporin or dermorphin) groups (n=3-6). The lesion of the nucleus raphe magnus (NRM) (n=3) only slightly reduced hyperalgesia at 3 h after inflammation. The loss of NGC mu-opioid receptor-containing neurons also decreased nocifensive behaviors only in phase II of the formalin model. In contrast, NRM lesions were without an effect on formalin-induced phase I/II responses. These findings indicate that selective deletion of the mu-opioid receptor-containing neurons in the nucleus reticularis giangocellularis attenuates inflammatory hyperagesia and allodynia.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Ventral tegmental area lesions alter EEG power spectrum across the sleep/wake cycle

Lee R, Gallegos RA, Crawford EF, Wills DN, Zhukov VI, Huitron-Resendiz S, Criado JR, Henriksen SJ (2003) Ventral tegmental area lesions alter EEG power spectrum across the sleep/wake cycle. Neuroscience 2003 Abstracts 616.5. Society for Neuroscience, New Orleans, LA.

Summary: The ventral tegmental area (VTA) has long been implicated in reward and drug abuse. We previously demonstrated (Lee et al, J. Neurosci. 2001) a role for VTA GABAergic neurotransmission in REM sleep. In continuing studies the potential role of the VTA in modulating electroencephalogram (EEG) activation was explored by selectively lesioning mu-opioid receptor expressing cells, or NMDA-lesioning cells, in the VTA. Under sodium pentobarbital anesthesia rats received either (1) a sham operation (2) a saporin injection (3) an injection of a dermorphin-saporin (DERM-SAP) conjugate (Advanced Targeting Systems, San Diego, CA) (4) or a bilateral VTA injection of NMDA. All injections were delivered in a volume of 0.5 to 1.0 µL over 4 to 8 minutes. Animals were also implanted with electrodes for recording the EEG & EMG. The filtered EEG & EMG were recorded continuously for 24 hours beginning 21 days after surgery. Frequency analysis of the EEG in 15-sec epochs revealed differences in the distribution of relative power in the DERM-SAP or NMDA-lesioned animals, compared to controls. Higher frequency components (12-25 Hz) were reduced in DERM-SAP lesioned animals during waking and slow wave sleep. Histology demonstrated gliosis of GAD-stained neurons in the VTA 3 to 4 weeks after injection of DERM-SAP. These data suggest that long-projecting GABA neurons of the VTA have a desynchronizing influence on cortical EEG arousal mechanisms. This is supported by anatomical evidence of both direct and indirect non-thalamic GABAergic projections to widespread areas of cortex in the rodent. Supported by: DA08301 to SJH.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Selective destruction of MOR expressing dorsal horn neurons using intrathecal dermorphin-saporin.

Wiley RG, Miller SA, Kline IV RH (2003) Selective destruction of MOR expressing dorsal horn neurons using intrathecal dermorphin-saporin. Neuroscience 2003 Abstracts 174.15. Society for Neuroscience, New Orleans, LA.

Summary: Evidence suggests that the mu opiate receptor (MOR) is key to the analgesic action of morphine. In the present study, we sought to determine if a disulfide conjugate of the mu opioid peptide, dermorphin, to the ribosome-inactivating protein, saporin, (derm-sap) would destroy neurons expressing MOR in the substantia gelatinosa (SG) of the spinal cord. Derm-sap was injected into the lumbar subarachnoid space of anesthetized adult, male Sprague-Dawley rats using a catheter inserted through the atlanto-occipital membrane and passed 8 cm caudally. The catheter was removed 15 minutes after toxin injection. Rats were sacrificed after 2 weeks, and 40 um transverse frozen sections of the L4 spinal segment were processed for immunohistochemical demonstration of MOR, NeuN, calbindin D28k, parvalbumin, NK-1R and for Nissl staining. In control rats, beta-funaltrexamine was injected just before derm-sap or derm-sap was pre-treated to reduce the disulfide bond which dissociates the toxin and neuropeptide. MOR staining in the SG was evaluated using quantitative densitometry. Initial experiments revealed a dose-related decrease in MOR staining in the dorsal horn without effect on dorsal root ganglia at doses up to 1000 ng. The maximally tolerated dose of derm-sap (500 ng) selectively decreased MOR staining by 54% as did multilevel lumbar dorsal rhizotomy. Combining 500 ng of derm-sap and multilevel rhizotomy produced 92% loss of MOR staining in the SG. Based on analysis of non-co-localized markers and control experiments, we interpret the results to indicate that intrathecal derm-sap selectively destroys MOR-expressing neurons in the SG without toxicity to primary afferents. This lesion will be useful in analysis of opioid mechanisms in the dorsal horn.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Selective elimination of mu-opioid receptor expressing neurons in the rostral ventromedial medulla (RVM) does not affect periaqueductal gray (pag) stimulation-produced analgesia

Harasawa I, Lai J, Porreca F, Fields HL, Meng ID (2003) Selective elimination of mu-opioid receptor expressing neurons in the rostral ventromedial medulla (RVM) does not affect periaqueductal gray (pag) stimulation-produced analgesia. Neuroscience 2003 Abstracts 177.5. Society for Neuroscience, New Orleans, LA.

Summary: PAG stimulation produces antinociception at spinal levels by modulating RVM neuronal activity. Microinjection of saporin conjugated with the mu-opioid receptor agonist dermorphin (DERM-SAP) into the RVM selectively eliminates MOR expressing neurons and diminishes neuropathic pain symptoms (Porreca et al., 2001). The aim of the present study was to determine whether MOR expressing neurons in the RVM are required for PAG stimulation produced analgesia (PAG/SPA). The minimum electrical current required to inhibit the tail flick response was compared in barbiturate-anesthetized rats given a single RVM injection of SAP or DERM-SAP 3-4 weeks prior to testing. Thresholds in SAP and DERM-SAP treated rats were not different. Furthermore, microinjection of the glutamate receptor antagonist kynurenic acid (10 mM, 800 nl) into the RVM disrupted PAG/SPA in both SAP and DERM-SAP treated rats. These results indicate that 1) mu-receptor expressing neurons in the RVM are not necessary for PAG/SPA, and 2) excitatory amino acid transmission in the RVM is critical for PAG/SPA. In additional experiments, inhibition of neurotransmitter release in the RVM by the microinjection of cobalt chloride (CoCl2, 100 mM, 800 nl), produced significant antinociception only in DERM-SAP treated rats. This finding suggests that DERM-SAP injections result in increased tonic inhibition of RVM neurons and that CoCl2 disinhibits these neurons to produce antinociception. Tonic inhibition of off-cells would account for our failure to find off-cells in DERM-SAP treated rats.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Enhanced evoked excitatory transmitter release in experimental neuropathy requires descending facilitation.

Gardell LR, Vanderah TW, Gardell SE, Wang R, Ossipov MH, Lai J, Porreca F (2003) Enhanced evoked excitatory transmitter release in experimental neuropathy requires descending facilitation. J Neurosci 23(23):8370-8379. doi: 10.1523/JNEUROSCI.23-23-08370.2003

Summary: The authors examine whether afferent discharge produced by nerve injury and central changes in experimental neuropathic pain might interact at the spinal level. Rats were treated with 48 ng of dermorphin-SAP (Cat. #IT-12) and various markers for neuropathic pain were evaluated. The results link several consequences of the post-injury state, including support for increased afferent input as a driving force for neuropathic pain.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Targeted toxins in pain.

Wiley RG, Lappi DA (2003) Targeted toxins in pain. Adv Drug Deliv Rev 55(8):1043-1054. doi: 10.1016/s0169-409x(03)00102-9

Summary: The authors discuss the use of 'molecular neurosurgery' in the study of nociception. Applications using targeted toxins, which include immunotoxins, protein-toxin conjugates, or peptide-toxin conjugates, are illustrated. The authors describe the use of these molecules as research tools, as well as their potential for therapeutics. A helpful table is included that lists neuronal surface markers and class of cells targeted for each targeted toxin. Reagents discussed: CTB-SAP (Cat. #IT-14), IB4-SAP (Cat. #IT-10), OX7-SAP (Cat. #IT-02), 192-Saporin (Cat. #IT-01), ME20.4-SAP (Cat. #IT-15), Anti-DBH-SAP (Cat. #IT-03), Anti-DAT-SAP (Cat. #IT-25), SP-SAP (Cat. #IT-07), Dermorphin-SAP (Cat. #IT-12), Orexin-SAP (Cat. #IT-20), CRF-SAP (Cat. #IT-13), and acetylated LDL-SAP (Cat. #IT-08).

Related Products: CTB-SAP (Cat. #IT-14), IB4-SAP (Cat. #IT-10), OX7-SAP (Cat. #IT-02), 192-IgG-SAP (Cat. #IT-01), ME20.4-SAP (Cat. #IT-15), Anti-DBH-SAP (Cat. #IT-03), Anti-DAT-SAP (Cat. #IT-25), SP-SAP (Cat. #IT-07), Dermorphin-SAP / MOR-SAP (Cat. #IT-12), Orexin-B-SAP (Cat. #IT-20), CRF-SAP (Cat. #IT-13), Acetylated LDL-SAP (Cat. #IT-08)

Selective lesion of ventral tegmental area neurons expressing mu-opioid receptors alters EEG power spectrum across sleep/wake cycle

Gallegos RA, Lee RS, Crawford E, Wills DN, Carr JR, Zhukov VI, Slaght KE, Huitron-Resendiz S, Criado JR, Henriksen SJ (2002) Selective lesion of ventral tegmental area neurons expressing mu-opioid receptors alters EEG power spectrum across sleep/wake cycle. Neuroscience 2002 Abstracts 276.14. Society for Neuroscience, Orlando, FL.

Summary: The ventral tegmental area (VTA) has long been implicated in motivated behaviors. Our previous study (Lee et al, J Neurosci 2001) also suggests a role for VTA GABAergic neurotransmission in REM sleep. In the current study the potential role of the VTA in modulating electroencephalogram (EEG) activation was explored by selectively deactivating mu-opioid receptor expressing cells in the VTA. Under sodium pentobarbital anesthesia, rats received either (1) a sham operation (2) a single bilateral VTA injection of NMDA (3) a saporin injection or (4) an injection of a dermorphin-saporin (DERM-SAP) conjugate (Advanced Targeting Systems, San Diego). Animals were also fitted with skull electrodes for recording the EEG. The filtered EEG was recorded continuously for 24 hours beginning 21 to 28 days after surgery. Frequency analysis of the EEG in 15-sec epochs revealed differences in the distribution of relative power in the DERM-SAP animals, compared to controls. Low frequency components (0.5-3.0 Hz and 3.0-8.0 Hz) were enhanced in dual lesioned animals during the dark phase but only during sleep. These results indicate that a selective inactivation of cells in the VTA has specific effects on arousal mechanisms in the intact animal.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Dermorphin-saporin targets descending facilitation in the rostral ventromedial medulla (rvm) to block CCK-induced abnormal pain

Burgess SE, Gardell LR, Xie Y, Ossipov MH, Vanderah TW, Malan TP, Porreca F, Lai J (2002) Dermorphin-saporin targets descending facilitation in the rostral ventromedial medulla (rvm) to block CCK-induced abnormal pain. Neuroscience 2002 Abstracts 351.11. Society for Neuroscience, Orlando, FL.

Summary: Abnormal pain from L5/L6 spinal nerve ligation (SNL) has been shown to require a time-dependent activation of descending facilitatory pathways arising in the RVM. Additionally, RVM microinjection of L365,260, a cholecystokinin (CCKB) receptor antagonist, reverses SNL-induced tactile and thermal hyperalgesia. These observations suggest the possibility that RVM CCK might “drive” such facilitation from the RVM by activating the endogenous descending facilitation system. Rats were treated with a single RVM injection of dermorphin (DERM) (μ opioid agonist), unconjugated saporin (SAP), or dermorphin-saporin (DERM-SAP) and responses to non-noxious tactile (von Frey filaments) or noxious radiant heat stimuli applied to the hindpaw were measured before and after RVM microinjection of CCK to uninjured rats. RVM DERM-SAP, DERM or SAP did not significantly alter baseline sensory thresholds over 28 days post-injection. At day 28, the rats received bilateral microinjections of CCK (30ng) in the RVM. Rats pretreated with DERM or SAP showed a time-related and revsersible CCK-induced tactile and thermal hypersensitivity. In contrast, RVM CCK failed to produce changes in sensory threshold in animals pretreated with DERM-SAP. The RVM pretreatments did not alter responses in control rats challenged with CCK vehicle. Additionally, lesions of the dorsolateral funiculus also blocked RVM CCK-induced tactile and thermal hypersensitivity. These data support the possibility of CCK-mediated activation of descending facilitation from the RVM as a mechanism of neuropathic pain.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Changes in rostral ventromedial medulla (RVM) neurons after the selective loss of mu-opioid receptor expressing cells.

Meng ID, Harasawa I, Lai J, Porreca F, Fields HL (2002) Changes in rostral ventromedial medulla (RVM) neurons after the selective loss of mu-opioid receptor expressing cells. Neuroscience 2002 Abstracts 351.9. Society for Neuroscience, Orlando, FL.

Summary: Different subpopulations of RVM neurons inhibit or facilitate dorsal horn nociceptive transmission. Microinjection of saporin conjugated to the mu-opioid receptor (MOR) agonist dermorphin (derm-sap) into the RVM selectively ablates MOR expressing neurons and diminishes neuropathic pain symptoms (Porreca et al., 2001). We examined the properties of neurons surviving a single RVM injection of derm-sap or sap control. Three classes of RVM neurons (On, Off, and Neutral) have been described with distinct responses to noxious stimuli and MOR agonists. On-cells increase and Off-cells cease firing just prior to a tail flick; MOR agonists inhibit On-cells and disinhibit Off-cells. Neutral cells are unaffected by either noxious stimulation or MOR agonists. Using single unit recording in lightly anesthetized rats a total of 10 electrode tracks were made per rat and each unit encountered was characterized according to its tail flick related activity. Injection of derm-sap (n=8) resulted in fewer On- and Off-cells when compared to saporin controls (n=8). The number of Neutral cells remained unchanged. In separate experiments, after derm-sap pretreatment RVM injections of the MOR agonist DAMGO were ineffective whereas injections of the glutamate receptor agonist homocysteic acid into the same sites increased tail flick latencies. The decrease in number of On-cells after derm-sap is consistent with evidence that these neurons express MOR and facilitate nociceptive transmission. The decrease in number of Off-cells indicates that inhibitory neurons responsible for producing the Off-cell tail flick related pause also express MOR.

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Enhanced morphine analgeisa after spinal dermorphin-saporin

Miller SA, Lappi DA, Wiley RG (2002) Enhanced morphine analgeisa after spinal dermorphin-saporin. Neuroscience 2002 Abstracts 218.1. Society for Neuroscience, Orlando, FL.

Summary: Dermorphin-saporin (derm-sap) is a neuropeptide toxin conjugate which is selective for neurons expressing the mu-opiate receptor (MOR). The dermorphin moiety of the conjugate binds MOR which is then internalized by the neuron, carrying the toxin with it. The saproin moiety inactivates ribosomes resulting in cell death. In the present study we sought to determine the effect of destroying MOR expressing neurons in Lamina II of the spinal cord dorsal horn on baseline thermal pain sensitivity and response to systemic morphine analgesia. 456 ng derm-sap (n=8) and vehicle (n=8) were injected into the lumbar CSF of adult male Sprague Dawley rats using a subarachnoid catheter inserted through the atlanto-occipital membrane and passed cadually to the level of the lumbar enlargement. 10 minutes following toxin injection, the catheters were withdrawn and the animals allowed to recover. When tested on a hotplate at 52C and on tail-flick assay, toxin rats did not differ from rats injected with vehicle. However, the dose-response curves for subcutaneous morphine were significantly shifted to the left (increased potency) in the toxin treated rats when compared with vehicle controls. Histological analysis of multiple dorsal root ganglia failed to reveal evidence of any primary afferent cell loss. We interpreted these findings to indicate that the neurons destroyed by derm-sap are lamina II MOR expressing neurons and play a role in morphine analgesia at high stimulus intensities.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Efferent projections from the striatal patch compartment: anterograde degeneration after selective ablation of neurons expressing mu-opioid receptor in rats.

Tokuno H, Chiken S, Kametani K, Moriizumi T (2002) Efferent projections from the striatal patch compartment: anterograde degeneration after selective ablation of neurons expressing mu-opioid receptor in rats. Neurosci Lett 332(1):5-8. doi: 10.1016/s0304-3940(02)00837-6

Summary: Taking advantage of the fact that neurons in patch compartments of the striatum express µ-opioid receptors, the authors injected 8.5 ng of dermorphin-SAP (Cat. #IT-12) into the striatum of rats. This lesion produced a degeneration of patch neurons as well as anterograde degeneration of efferent fibers from patch compartments, allowing further elucidation of the functional organization of the striatum.

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Descending facilitation from the rostral ventromedial medulla maintains, but does not initiate, neuropathic pain.

Burgess SE, Gardell LR, Ossipov MH, Malan T, Vanderah TW, Lai J, Porreca F (2002) Descending facilitation from the rostral ventromedial medulla maintains, but does not initiate, neuropathic pain. IASP 2002 Abstracts International Association for the Study of Pain, San Diego, CA.

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Chronic pain and medullary descending facilitation

Porreca F, Ossipov MH, Gebhart GF (2002) Chronic pain and medullary descending facilitation. Trends Neurosci 25(6):319-325. doi: 10.1016/s0166-2236(02)02157-4

Objective: To examine the likelihood that sustained activation of descending modulatory pathways that facilitate pain transmission could underlie some states of chronic pain.

Summary: Rats treated with Dermorphin-SAP, either before or after spinal nerve ligation injury, did not display neuropathic pain behaviors, although normal nociceptive responses were intact.

Usage: Rostroventromedial medulla (RVM) injected with Dermorphin-SAP.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

See Also:

Time-dependent descending facilitation from the rostral ventromedial medulla maintains, but does not initiate, neuropathic pain.

Burgess SE, Gardell LR, Ossipov MH, Malan Jr TP, Vanderah TW, Lai J, Porreca F (2002) Time-dependent descending facilitation from the rostral ventromedial medulla maintains, but does not initiate, neuropathic pain. J Neurosci 22(12):5129-5136. doi: 10.1523/JNEUROSCI.22-12-05129.2002

Summary: Various indications, such as declining afferent discharge over time, suggest that the mechanisms involved in persistent neuropathic pain are different than those that initiate the pain. The authors have previously shown that cells expressing the mu-opioid receptor are involved in the descending pain pathway. In this work, the authors lesioned the rostral ventromedial medulla (RVM) in rats using 1.5 pmol in 0.5 µl of dermorphin-SAP (Cat. #IT-12) administered to each side of the RVM. Measurements of pain-related behavior show that mu-opioid receptor-expressing cells in the RVM are involved in the maintenance of heightened sensitivity to stimuli seen in neuropathic pain.

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Dermorphin-saporin conjugate relieves inflammatory pain after peripheral application.

Palecek J, Paleckova V, Willis WD (2001) Dermorphin-saporin conjugate relieves inflammatory pain after peripheral application. Neuroscience 2001 Abstracts 508.10. Society for Neuroscience, San Diego, CA.

Summary: Opioid receptors have been shown to exist in specific population of DRG neurons signaling nociceptive information from peripheral tissues. In our study, we attempted to selectively destroy these neurons by using a peripheral application of the mu opioid agonist Dermorphin conjugated to ribosome inactivating toxin Saporin (DERM-SAP, Advanced Targeting Systems) in order to alleviate inflammatory pain. Intraarticular or intraplantar injection of carrageenan or CFA was used to induce inflammation in rats. The DERM-SAP conjugate was injected into the inflamed area 12-48h later. Responses of the animals to mechanical and thermal stimuli were tested before and after the inflammation and up to 21 days after the DERM-SAP application. The rats developed heat hyperalgesia in the affected paw 24 hours after the intraarticular CFA injection. In the saline injected group the hyperalgesia persisted for up to 19 days, but in the DERM-SAP injected group the signs of hyperalgesia were improving from day 7. Also mechanical allodynia tested with a VF filament (1.1g) was alleviated in the DERM-SAP group. In the carrageenan group, the DERM-SAP treatment decreased the heat hyperalgesia and prevented the development of hyperalgesia after repeated carrageenan application, 21days after the DERM-SAP treatment. Postmortem evaluation with a specific antibody showed presence of saporin in the DRG neurons. Our results show that peripheral application of DERM-SAP relieves inflammatory pain and suggest that peripheral application of neuropeptides conjugated to cell toxins or other substances such as antisense probes could be a useful tool for treating pain of peripheral origin. Supported by NIH grants NS09743 and NS11253.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Inhibition of neuropathic pain by selective ablation of brainstem medullary cells expressing the µ-opioid receptor.

Porreca F, Burgess SE, Gardell LR, Vanderah TW, Malan TP Jr, Ossipov MH, Lappi DA, Lai J (2001) Inhibition of neuropathic pain by selective ablation of brainstem medullary cells expressing the µ-opioid receptor. J Neurosci 21(14):5281-5288. doi: 10.1523/JNEUROSCI.21-14-05281.2001

Summary: The presence of descending projections in the pain pathway raises the possibility that abnormal sustained activity may perpetuate chronic pain. Using 3-ng injections of dermorphin-SAP (Cat #IT-12) on either side of the RVM in rats the authors both prevented and reversed neuropathic pain caused by spinal nerve ligation.

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Featured Article: Dermorphin-SAP kills MOR-positive cells

Lappi DA (2001) Featured Article: Dermorphin-SAP kills MOR-positive cells. Targeting Trends 2(3)

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Read the featured article in Targeting Trends.

Intrathecal dermorphin-saporin decreases morphine effect in hotplate algesia testing.

Miller SA, Lappi DA, Wiley RG (2000) Intrathecal dermorphin-saporin decreases morphine effect in hotplate algesia testing. Neuroscience 2000 Abstracts 212.8. Society for Neuroscience, New Orleans, LA.

Summary: The targeted cytotoxin, dermorphin-saporin, selectively destroys cells expressing MOR. In the present study, we gave dermorphin-saporin by lumbar i.t. injection and sought to determine if destroying dorsal horn neurons expressing MOR would alter thermal sensitivity and/or response to systemic morphine (MS) using hotplate testing under various conditions. 16 male Sprague-Dawley rats were tested on constant temperature (0.3, 44 and 47 C) and incremental (0.1 C/sec from 28 to 57 C) hotplates. Then 8 rats received lumbar intrathecal injections of derm-sap (465 ng) and 8 received vehicle using a subarachnoid PE-10 catheter that was removed 10 mins after injection. Retesting rats after toxin/vehicle injection showed no change in responses to any of the hotplate conditions. However, vehicle but not derm-sap rats showed increased lick latency on the incremental hotplate 20 mins after MS, 2.5 mg/kg, s.c. At 5 mg/kg of MS, vehicle and dermorphin-saporin rats showed identical responses. Capsaicin cream (0.94%) applied to the plantar surface of both hindpaws 3 hrs before testing on the 44 C hotplate produced decreased lick latencies in both groups of rats. MS, 5 mg/kg, s.c., produced increased lick latencies in capsaicin treated vehicle but not derm-sap rats. At 10 mg/kg, MS produced identical effects in capsaicin treated vehicle and toxin rats. These results indicate that i.t. derm-sap produced no change in baseline thermal sensitivity but did diminish the effect of low dose MS under conditions that preferentially test C nociceptor function suggesting that MOR-expressing dorsal horn neurons play a role in the analgesic action of low dose MS.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Dermorphin-saporin targets tonic descending facilitation in the rostral ventromedial medulla to block and reverse neuropathic pain.

Burgess SE, Vanderah TW, Mantyh PW, Malan Jr TP, Ossipov MH, Lappi D, Lai J, Porreca F (2000) Dermorphin-saporin targets tonic descending facilitation in the rostral ventromedial medulla to block and reverse neuropathic pain. Neuroscience 2000 Abstracts 243.6. Society for Neuroscience, New Orleans, LA.

Summary: The hypothesis that chronic pain from L5/L6 spinal nerve ligation (SNL) is due to tonic activation of descending pain facilitation mechanisms was explored by selective targeting mu (μ) opioid receptor expressing cells in the RVM (i.e., presumably, ON cells). Rats were treated with a single RVM injection of dermorphin (DERM)(μ agonist), saporin (SAP), or dermorphin-saporin conjugate (DERM-SAP) and responses to non-noxious (von Frey filaments) or noxious (Hargreave's test) stimuli characterized. DERM-SAP retained high affinity for μ receptors and acutely produced antinociception (tail-flick test), indicating agonist actions of the conjugate. Decreased staining of μ receptor-expressing cells was seen in superficial dorsal horn and in dorsal root ganglia 28 days after intrathecal injection of DERM-SAP, but not DERM or SAP. RVM DERM-SAP, DERM or SAP did not significantly alter baseline thresholds to von Frey filaments or noxious heat applied to the paw over 28 days. At day 28, RVM pretreated rats were subjected to sham- or SNL surgery and responses to tactile and heat stimuli monitored 7 days later (i.e., 35 days after the RVM pretreatment). DERM and SAP pretreated SNL rats showed the expected development of tactile allodynia and thermal hyperalgesia, while DERM-SAP pretreated rats did not. The RVM pretreatments did not alter responses in sham-operated controls. Administration of RVM DERM-SAP, but not SAP or DERM, to SNL rats showed full reversal of established allodynia/hyperalgesia by day 14. RVM pretreatment with β-funaltrexamine (β-FNA; opioid μ antagonist) prevented the antiallodynic and antihyperalgesic effects of subsequent DERM-SAP injection. These data, together with findings of blockade of SNL pain with RVM lidocaine or lesions of the dorsolateral funiculus, support the possibility of tonic activation of descending facilitation as a basis for chronic neuropathic pain.

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Entering through the doors of perception: characterization of a highly selective Substance P receptor-targeted toxin.

Lappi DA, Wiley RG (2000) Entering through the doors of perception: characterization of a highly selective Substance P receptor-targeted toxin. Neuropeptides 34(5):323-328. doi: 10.1054/npep.2000.0827

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Synthesis and receptor binding analysis of dermorphin hepta-, hexa- and pentapeptides.

Attila M, Salvadori S, Balboni G, Bryant SD, Lazarus LH (1993) Synthesis and receptor binding analysis of dermorphin hepta-, hexa- and pentapeptides. Int J Pept Prot Res 42:550-559. doi: 10.1111/j.1399-3011.1993.tb00363.x

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)

Amino acid composition and sequence of dermorphin, a novel opiate-like peptide from the skin of Phyllomedusa sauvagei.

Montecucchi PC, de Castiglione R, Piani S, Gozzini L, Erspamer V (1981) Amino acid composition and sequence of dermorphin, a novel opiate-like peptide from the skin of Phyllomedusa sauvagei. Int J Pept Prot Res 17(3):275-283. doi: 10.1111/j.1399-3011.1981.tb01993.x

Related Products: Dermorphin-SAP / MOR-SAP (Cat. #IT-12)