References

Abudukeyoumu N, Garcia-Munoz M, Nakano Y, Arbuthnott GW (2018) Impaired reach-to-grasp responses in mice depleted of striatal cholinergic interneurons. Neuroscience 2018 Abstracts 491.01 / MM13. Society for Neuroscience, San Diego, CA.

Summary: Cholinergic interneurons (ChIs) are sparsely distributed within the striatum, a nucleus that plays important role in voluntary motor control, associated learning, procedural memory, action selection and planning and execution of movement. Sparsely distributed ChIs are 1-3% of all striatal neurons and the main source of striatal acetylcholine. Here we report the effect of depletion of ChIs in the dorsolateral striatum in a reach-to-grasp task. To selectively deplete ChIs, we used the saporin ribosome-inactivating-immunotoxin that targets choline acetyltransferase. C57BL/J male mice, 21 days old, received a stereotaxic unilateral infusion of the toxin (0.3µl/3min), and sham control group was injected with saline. Following one week postsurgery recovery, animals were food deprived for 12 h everyday and trained for 12 days at night during their active circadian cycle. The mean percentage ± SEM of successful performance in the reach-to-grasp task for the last 6 training sessions was 51.11 ± 4.09% (n = 25), 48.79 ± 7.7% (n = 9) and 26.28 ± 5.19% (n = 13) for intact control, sham control and ChIs-depleted mice, respectively. These results indicate that striatal depletion of ChIs impair reaching accuracy, whereas no significant differences were observed in control or sham operated mice. Moreover, a positive correlation between loss of ChIs and performance in the reach-to-grasp task was observed. Our results suggest that the participation of ChIs in striatal mediated motor learning impact on the function of interneurons and projection neurons of the whole striatal microcircuitry (Abudukeyoumu, N., Hernandez-Flores, T. et al. Eur. J. Neuroscience, in press).

Related Products: Anti-ChAT-SAP (Cat. #IT-42)

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Gulisano M, Vicario N, Costantino A, Giunta MAS, Spitale FM, Parenti R, Gulino R (2018) Sonic hedgehog signalling pathway during regenerative processes in a mouse model of spinal motoneuronal loss. Neuroscience 2018 Abstracts 379.29 / L1. Society for Neuroscience, San Diego, CA.

Summary: Background – Neuronal loss represents the consequence of direct or indirect insults to neurons, as well as one of the major factors mediating persistent disability. Gliosis, neuroinflammation and neurodegeneration are processes in which different cell populations have an interplay mediating both hostile microenvironment and self-repairing mechanisms. Sonic hedgehog (Shh) signaling, which has been indicated as an important pathway in central nervous system development and neural stem cells (NSCs) function, may have a role in prompting the repairing and modulating actions of endogenous and/or exogenous NSCs in neurodegenerative conditions. Methodology – Somatic NSCs were obtained from the subventricular zone (SVZ) of 8 week old female 129/Sv mice. We studied the Shh pathway on NSCs both in vitro and in a mouse model of spinal motorneuronal degeneration induced by cholera toxin B conjugated to saporin (CTB-Sap) injection into the gastrocnemius muscle. Results – NSCs were derived, expanded and characterized in vitro. We analyzed the effects of Shh signaling pathway modulation on NSCs in vitro, finding a significant increase of the NSCs growth rate (2.98±0.58 vs. 5.26±0.57, p<0.05) and neurospheres diameters (109.9±2.4μm vs. 129.6±3.7μm, p<0.01) upon Shh pathway activation. We then characterized Shh signaling activation in CTB-Sap mice analyzing neuronal loss, gliosis, inflammation and compensatory self-repairing mechanisms, compared to intact control mice. Conclusions - Our results suggest a crucial role of Shh signaling during regenerative processes and NSCs as a potential strategy to support recovery after spinal motoneuronal degeneration, thus representing a promising approach for neurodegenerative disorders.

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Cammarata C, DeRosa ED, Anderson AK (2018) Lifespan and cholinergic changes in cognitive flexibility in rats. Neuroscience 2018 Abstracts 512.05 / GGG8. Society for Neuroscience, San Diego, CA.

Summary: The ability to update one’s mental schemas in order respond flexibly and adaptably – i.e. cognitive flexibility – is crucial to navigating a dynamic environment. Proactive interference (PI) is a phenomenon wherein prior memory impedes the formation of new memories for similar information, biasing behavior toward no-longer-relevant schemas. Thus, overcoming PI is an important aspect of cognitive flexibility. PI is exacerbated during aging, and in turn contributes to age-related deficits in cognitive flexibility. In young animals and young adult humans, resolution of PI has been found to rely on neuromodulatory activity via Acetylcholine (ACh), and ACh levels are known to decline in aging, however it has yet to be demonstrated whether these age-related changes in ACh directly contribute to age-related increase in PI. Here, we first compared PI resolution in middle-aged (13 months, n = 8) and old (23 months, n= 11) male Long Evans rats, finding that old animals were more inefficient in resolving PI when compared to the middle-aged animals. Furthermore we performed cholinergic deafferentation, with the immunotoxin 192-IgG saporin (SAP; 0.2 µl of 0.3 µg/µl dissolved in sterile phosphate buffered sale in each of four locations targeting bilateral anterior and posterior basal forebrain), in our older rats (N= 5 SAP and N=6 Sham) which had no effect on the floor performance of older rats. This suggests that the inability to resolve PI seen in the aged rats may be due to already-depleted levels of ACh. We are currently collecting local field potential data in the prelimbic and posterior parietal cortices in behaving older and younger rats and will combine this with central administration of muscarinic cholinergic pharmacology to continue to examine age-related changes in the cortical dynamics that support cognitive flexibility. Based on prior findings in our laboratory examining similar attentional flexibility, we predict the young animals will demonstrate increased beta band LFP activity in the posterior parietal cortex, and potentially increased beta coherence between prefrontal and posterior parietal cortices, related to successful resolution of PI. We expect such activity to be mitigated by cholinergic antagonists and in the older animals.

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Daripelli S, Bhayrapuneni G, Tirumalesetty C, Benade V, Subramanian R, Petlu S, Praveena N, Jayarajan P, Shinde A, Badange R, Bhatta V, Nirogi R (2018) SUVN-G3031, H3 receptor inverse agonist produces wake promoting activity in rats with hypocretin-2-saporin lesions of the lateral hypothalamus. Neuroscience 2018 Abstracts 679.23 / VV4. Society for Neuroscience, San Diego, CA.

Summary: Numerous studies have demonstrated that brain histamine plays a crucial role in maintenance of wakefulness, attention, learning and other cognitive processes. SUVN-G3031, a potent H3 receptor inverse agonist is being developed for the treatment of narcolepsy and other sleep related disorders. SUVN-G3031 is one of the lead molecules with hKi of 8.7 nM and has more than 100 fold selectivity against the related GPCRs. SUVN-G3031 exhibited desired pharmacokinetic properties and brain penetration. SUVN-G3031 blocked R-α-methylhistamine induced water intake and increased tele-methylhistamine levels in brain and cerebrospinal fluid. A single oral administration of SUVN-G3031 produced significant increase in acetylcholine, histamine, dopamine and norepinephrine levels in the cortex. SUVN-G3031 produced wake promoting activity in male Wistar rats. In the present study, effects of SUVN-G3031 on sleep/ wake profile were evaluated in rats with lateral hypothalamic lesion using neurotoxin hypocretin-2-saporin. Narcoleptic-like sleep behavior was observed in rats injected with hypocretin-2-saporin in lateral hypothalamus. SUVN-G3031 produced significant increase in wakefulness with concomitant decrease in rapid eye movement (REM) sleep in these animals. These results are in agreement with electroencephalography (EEG) studies carried out in healthy male Wistar rats. Results from the current study and the neurotransmitter modulations produced by SUVN-G3031 provide a strong basis for the potential of SUVN-G3031 in treatment of sleep related disorders. First in human, Phase 1 studies for SUVN-G3031 are completed underUS IND and SUVN-G3031 has shown desirable pharmacokinetic profile with safety and tolerability in healthy human volunteers. Phase 2 study for narcolepsy is currently being planned.

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Chew C, Sengelaub DR (2018) Exercise is neuroprotective following partial motoneuron depletion: Run for your dendrites. Neuroscience 2018 Abstracts 761.02 / MM11. Society for Neuroscience, San Diego, CA.

Summary: We have previously demonstrated that partial depletion of motoneurons innervating the quadriceps muscles induces dendritic atrophy in remaining motoneurons. Furthermore, systemic treatment with supplemental androgens is neuroprotective and dendritic atrophy following partial motoneuron depletion is attenuated. Circulating levels of androgens have previously been shown to increase following exercise, and exercise has been demonstrated to be neuroprotective in a variety of other neurodegenerative and injury models. Thus, we hypothesized that allowing animals to exercise following partial motoneuron depletion would produce neuroprotective effects similar to treatment with supplemental androgens. Motoneurons innervating the vastus medialis muscle in adult male rats were selectively killed by intramuscular injection of cholera toxin-conjugated saporin. Following saporin injections, some animals were allowed free access to a running wheel attached to their home cages. Four weeks later, motoneurons innervating the ipsilateral vastus lateralis muscle were labeled with cholera toxin-conjugated horseradish peroxidase, and dendritic arbors were reconstructed in three dimensions. Compared with intact normal males, partial motoneuron depletion resulted in decreased dendritic length in remaining quadriceps motoneurons. Early data suggests that exercise can completely protect against this dendritic atrophy, with exercised males showing dendritic arbors lengths significantly longer than saporin and testosterone-treated animals, and of similar length to intact normal animals. These findings suggest that exercise may be a viable means of protecting against collateral dendritic atrophy. The upregulation of testosterone release following exercise combined with our previous data showing the neuroprotective effects of androgen treatment suggest that the neuroprotective following exercise may be attributable to systemic androgen upregulation.

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Goodman RL, Lopez JA, Bedenbaugh MN, Connors JM, Hardy SL< Hileman SM, Coolen LM, Lehman MN (2018) Evidence that the LH surge in ewes involves both neurokinin B-dependent and -independent actions of kisspeptin. Neuroscience 2018 Abstracts 773.20 / YY14. Society for Neuroscience, San Diego, CA.

Summary: It is generally recognized that kisspeptin plays a key role in induction of the LH surge in sheep and we have reported evidence that neurokinin B (NKB) does so as well. Specifically, disrupting NKB signaling in the retrochiasmatic area (RCh) using either an antagonist to its receptor, NK3R, or lesions of NK3R-containing neurons in the RCh with a saporin conjugate (NK3-SAP) reduced the amplitude of the estrogen-induced LH surge by 50%. Because a KISS1R antagonist (p271) also produced a 50% decrease in surge amplitude, we hypothesized that these two systems are organized in series with NKB actions in the RCh stimulating kisspeptin release. If this is the case, then the combination of NK3R lesions and a KISS1R antagonist should produce the same inhibition as either treatment alone. This experiment tested this prediction using a 2 x 2 design. Breeding season ewes were ovariectomized and immediately given an estradiol (E) implant sc and two progesterone implants (CIDRs) intravaginally that produced luteal phase levels of these steroids. Ewes then received bilateral injections of either NK3-SAP (n=6) or Blank-SAP (n=5) into the RCh. Three weeks later, an artificial follicular phase was produced by inserting four 3 cm long E implants 24 hrs after CIDR removal and either saline or p271 was infused into the lateral ventricle for 16-24 hrs after E implantation; LH was monitored every 2-4 hrs for two days. CIDRs were then reinserted and the protocol repeated in a cross-over design so that all ewes received saline and p271 treatment. In Blank-SAP ewes, p271 decreased the peak of the LH surge from 61.2 ± 7.6 to 27.4 ± 4.6 ng/mL and delayed it 8 hrs (from 26.5 ± 0.5 to 34.1 ± 1.2 hrs post E implantation). The NK3-SAP injections alone decreased the peak of the LH surge to 29.7 ± 10.7 ng/mL compared to Blank-SAP, but the peak was not further inhibited by p271 in these NK3-SAP-treated ewes (24.4 ± 1.4 ng/mL). However, p271 delayed the peak of the LH surge (from 28.8 ± 1.2 to 34.8 ± 2.1 hrs post E implantation) in the ewes injected with NK3-SAP. Based on these results, we propose that kisspeptin has two roles in the LH surge in ewes: it initiates the surge independent of NKB signaling in the RCh, and maintains LH secretion during the surge by a NKB-dependent system.

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Moreno-Rodriguez M, Martinez-Gardeazabal J, Llorente-Ovejero A, Lombardero L, Manuel I, Rodriguez-Puertas R (2018) Learning and memory improvement mediated by CB1 cannabinoid receptors in animal models of cholinergic dysfunction. Neuroscience 2018 Abstracts 049.05 / S3. Society for Neuroscience, San Diego, CA.

Summary: The selective vulnerability of the basal forebrain cholinergic system (BFCS) is responsible for most of the clinical alterations in learning and memory processes that are characteristic of the Alzheimer’s disease (AD). The loss of cholinergic neurons and muscarinic receptors (MR) in the nucleus basalis of Meynert have been reported in AD. The endocannabinoid system is a neuromodulator of the BFCS, but there are controversial reports regarding the cannabinoid effects in learning and memory processes. The animal models of cholinergic impairment mimic the main histopathological and behavioral effects observed in patients. The MR antagonism, e.g. using scopolamine (SCOP), is used as a model of amnesia in rodents. The intraparenchymal administration of 192-IgG-saporin (SAP) in the nucleus basalis magnocellularis eliminates cholinergic neurons leading to learning and memory deficits. Then, the present study evaluates the modulation of spatial and working memory with the Barnes Maze following a subchronic treatment with a low dose (0.5 mg/kg) of WIN55,212-2 (WIN) in both the SCOP and SAP models of learning and memory deficit. In the SCOP model, the administration of WIN protects learning and memory impairment during the probe trial, recorded as the time spent in the target quadrant (WIN + SCOP: 78 ± 13 sec vs VEH + SCOP: 45 ± 3 sec; p < 0.001). A similar effect of the treatment was observed in the SAP model (SAP: 50 ± 3 sec vs SAP + WIN: 82 ± 7 sec; p < 0.001). This effect was specifically mediated by CB1 receptors, since it was blocked by the co-administration of the specific CB1 antagonist, SR141716A (0.5 mg/kg) (SAP: 49 ± 3 sec vs SAP + WIN + SR: 48 ± 5 sec). However, higher doses of WIN (3 mg/kg) induced negative effects in learning and memory in control (C) rats, but positive and comparable to the lower dose in the SAP model (C: 89 ± 3 sec, C + WIN-3 mg/kg: 48 ± 3 sec; SAP: 49 ± 3; SAP + WIN-3 mg/kg: 80 ± 12 sec; p < 0.001). The CB1 receptor activation by low doses of the cannabinoid agonist WIN are able to block the amnesic effects induced by SCOP and also the learning and memory impairment produced by the BFCS pathway degeneration. CB1 agonists could contribute to improve the clinical symptoms of AD. International application patent PCT/EP2018/054525.

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Zoccal DB, Silva JN, Barnett WH, Lemes EV, Falquetto B, Colombari E, Molkov YI, Moreira TS, Takakura AC (2018) Interaction between the retrotrapezoid nucleus and the parafacial respiratory group to regulate active expiration and sympathetic activity in rats. Am J Physiol Lung Cell Mol Physiol 315(5):L891-L909. doi: 10.1152/ajplung.00011.2018

Objective: To investigate the microcircuitry responsible for the distribution of the excitatory signals to the the parafacial respiratory group (pFRG) and the the respiratory central pattern generator (rCPG) in conditions of high CO2.

Summary: The activation of the pFRG late-E neurons during hypercapnia require glutamatergic inputs from the RTN neurons that intrinsically detect changes in CO2/pH.

Usage: Bilateral injections of SSP-SAP (0.6 ng in 100 nL of saline per side).

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Qian L, Milne MR, Shepheard S, Rogers ML, Medeiros R, Coulson EJ (2019) Removal of p75 neurotrophin receptor expression from cholinergic basal forebrain neurons reduces amyloid-β plaque deposition and cognitive impairment in aged APP/PS1 mice. Mol Neurobiol 56(7):4639-4652. doi: 10.1007/s12035-018-1404-2

Objective: To investigate the contribution of CBF neuronal p75NTR to the progression of Alzheimer’s Disease

Summary: Data indicate that a direct interaction between CBF-expressed p75NTR and Aβ does not contribute significantly to the regulation of Aβ load.

Usage: To lesion CBF neurons, a single infusion of mu p75-SAP or control Rabbit IgG-SAP (0.4 mg/ml) was stereotaxically-injected into the basal forebrain.

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Garrett AM, Khalil A, Walton DO, Burgess RW (2018) DSCAM promotes self-avoidance in the developing mouse retina by masking the functions of cadherin superfamily members. Proc Natl Acad Sci U S A 115:E10216-E10224. doi: 10.1073/pnas.1809430115

Summary: Focus on DSCAM (Down syndrome cell adhesion molecule 1) self-avoidance function in the mouse retina. DSCAM and members of the cadherin superfamily have also emerged as key contributors to a variety of neurodevelopmental disorders, including autism, schizophrenia, bipolar disease, Down syndrome and intellectual disability.

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