Substance P, NPY, CCK and their receptors in five brain regions in major depressive disorder with transcriptomic analysis of locus coeruleus neurons

Major depressive disorder (MDD) is a serious disease and a burden to patients, families and society. Rodent experiments and human studies suggest that several neuropeptide systems are involved in mood regulation. The aim of this study is two-fold: (i) to monitor, with qPCR, transcript levels of the substance P/tachykinin (TAC), NPY and CCK systems in bulk samples from control and suicide subjects, targeting five postmortem brain regions including locus coeruleus (LC); and (ii) to analyse expression of neuropeptide family transcripts in LC neurons of ‘normal ’ postmortem brains by using laser capture microdissection with Smart-Seq2 RNA sequencing. qPCR revealed distinct regional expression patterns in male and female controls with higher levels for the TAC system in the dorsal raphe nucleus and LC, versus higher transcripts levels of the NPY and CCK systems in prefrontal cortex. In suicide patients, TAC, TAC receptors and a few NPY family transcript levels were increased mainly in prefrontal cortex and LC. The second study on ‘normal ’ noradrenergic LC neurons revealed expression of transcripts for GAL, NPY, TAC1, CCK , and TACR1 and many other peptides (e.g. Cerebellin4 and CARTPT ) and receptors (e.g. Adcyap1R1 and GPR173 ). These data and our previous results on suicide brains indicates that the tachykinin and galanin systems may be valid targets for developing antidepressant medicines. Moreover, the perturbation of neuropeptide systems in MDD patients, and the detection of further neuropeptide and receptor transcripts in LC, shed new light on signalling in noradrenergic LC neurons and on mechanisms possibly associated with mood disorders.


Introduction
Major depressive disorder (MDD) is a serious disease, afflicting more women than men and is associated with much suffering and major costs for society (Wittchen, 2012;Murray et al., 2013;World Health Organization, 2017), and until recently often also with treatment resistance (McAllister-Williams et al., 2020).Interaction between genetic and environmental factors with stressful life events may represent important predisposing features (Nestler et al., 2002).Over decades, monoamines have been implicated in the processes that lead to the symptoms of MDD, both noradrenaline (NA) (Schatzberg and Schildkraut, 1995), and serotonin (Maes and Meltzer, 1995;Sharp and Cowen, 2011), a view supported by the antidepressant effects of drugs that enhance signalling of these monoamines (Millan, 2006).More recently a paradigm shift, from monoamines to amino acids, has been announced based on breakthrough research, showing that the glutamate receptor antagonist ketamine has a rapid effect on treatment-resistant depression (Kadriu et al., 2019) (Krystal et al., 2019), as have psychedelics (Johnston et al., 2023) and transcranial magnetic stimulation (Cole et al., 2020).Also neuropeptides, the most diverse group of transmitters (Burbach, 2010), have been considered as targets for treatment of stress-related disorders (Maubach et al., 1999;Holmes et al., 2003;Griebel and Holsboer, 2012;Brockway and Crowley, 2020;Casello et al., 2022;Kupcova et al., 2022;Singewald et al., 2023).For recent reviews on peptide signalling, see (Eiden et al., 2022;Girven et al., 2022).
Substance P (SP)/tachykinin (TAC), neuropeptide Y (NPY), cholecystokinin (CCK) and their multiple receptors are widely expressed and distributed in the rodent and human brain, as shown in a large number of studies using various methods.All these three systems have been analysed in extensive animal experiments regarding a possible role in various behaviors and pathological states, including mood derangements.Moreover, attempts have been made to explore a role and therapeutic potential of these peptide systems in human mood disorders, including a trial with an NK1 antagonist (Kramer et al., 1998;Keller et al., 2006).NK1 antagonists have also been studied in other contexts, like posttraumatic stress disorder (Frick et al., 2016).Regarding NPY a randomized, controlled trial of intranasal administration of NPY to subjects with MDD has been performed (Mathé et al., 2020).
We have previously reported on the expression of galanin and its three receptors, GALR1-3, in bulk samples from five regions of post-mortem brains (Fig. 1) from depressed subjects who died by suicide (DSS) and controls: dorsolateral prefrontal cortex (DLPFC), anterior cingulate cortex (ACC), the dorsal raphe nucleus (DRN), locus coeruleus (LC)] and medullary raphe nuclei (MRN) (Barde et al., 2016).In the present study, we have processed the same tissue samples using qPCR analysis, with focus on the SP/TAC, NPY and CCK systems.
Since these studies are based on micropunched bulk samples using qPCR, we do not know in which type of neurons our markers are present.We have therefore added a second study based on RNA sequencing analysis of single neurons (about 50-120) isolated and then pooled from the LC of 20 'normal' subjects using laser capture microdissection (LCM).This allows us to validate the results on the LC from our qPCR analysis at least in one region, that is to demonstrate that neuropeptide system markers analysed are indeed present in noradrenergic LC neurons.In addition, we could explore to what extent these LC neurons express more transcripts than the ones mentioned above.

qPCR analysis
Postmortem brain tissue was obtained from the Douglas-Bell Canada Brain Bank (Montreal, Canada).A total of 212 punched brain samples from five different regions were received (Table 1).Written informed consent for post-mortem brain donation was received from the donors' families, and the ethical approval for this study was obtained from the Institutional Review Board of the Karolinska Institutet (see SI Text and Table S1).

Laser capture microdissection
Fresh frozen postmortem brain tissue including the LC was obtained from the Netherland's Brain Bank (NBB) and NIH Neurobiobank.Samples from a total of 20 individuals were included (See Table S2).Ethical approval for this work was obtained from the regional ethical review board of Stockholm, Sweden (EPN Dnr 2012/111-31/1).The work with human tissues was carried out according to the Code of Ethics of the World Medical Association (Declaration of Helsinki).

RNA isolation, integrity analysis and quantitative real time PCR (qPCR)
RT-qPCR was performed as described previously (Le Maitre et al., 2013) with some modifications as described in detail in the Supplementary Information (SI).For details, see SI text.Fig. 1.Schematic overview of the five brain subregions used for analysis.Five subregions of the human brain were microdissected and used for qPCR analysis from male and female controls and depressed suicide subjects; these included, dorsolateral prefrontal cortex (DLPFC, BA8/9), anterior cingulate cortex (ACC, BA 24), dorsal raphe nucleus (DRN), locus coeruleus (LC) and medullary raphe nucleus (MRN).For the LCM-seq analysis, individual cells from the locus coeruleus were laser captured, and 50-120 cells were pooled and processed for transcriptomic sequencing.

Statistical analysis
Statistical analysis was performed with GraphPad Prism 6 (GraphPad software, CA) StatView (SAS Institute Inc.).The different variables: RIN, age, postmortem interval and tissue pH values between controls and suicides for the five brain regions were analysed by multivariate ANOVA and two-tailed t-test for independent groups.Shapiro-Wilks test was used to test for Gaussian distribution.Significant outliers for qPCR fold change were analysed by ROUT method and excluded if p < 0.05.qPCR fold change data was analysed by using the nonparametric Mann-Whitney U test.p-values below 0.05 were considered significant, and p-values between 0.05 and 0.1 were considered to represent a trend.

Laser capture microdissection, RNA seq and phenotyping of LC neurons
Briefly, 10 um thick coronal sections from human pons was used, and the LC neurons were selected based on the location and presence of neuromelanin.To secure detection of low abundance transcripts 50-120 neurons were pooled from each subject and a single cDNA library was prepared.This was followed by sequencing using either the HiSeq2500 or the NovaSeq platforms and determination of expression levels as normalized counts.A detailed description of this method can be found in the SI Text.

Cohort demographics and effects of medication
No significant differences (mean ± s.d.) between DSS and their matched controls were observed for age, postmortem interval, brain pH or RIN for qPCR and LCM-seq studies.Demographic characteristic details of each subject used for each of the regions analysed are provided in Table 1a and 1b, and detailed information on each individual subject is provided in Table S1 and S2.For detailed statistical analysis, see SI Text.
Similarly, to analyse if medications act as confounding factors which is an innate problem with autopsy studies, samples were divided into 4 groups based on the information of the medications received by the patients.This data was analysed using ANOVA followed by Fisher's LSD post-hoc tests and ANCOVA by using antidepressants as a confounding factor.No significant effects on the gene expressions of the peptide systems studied were recorded.For details see SI Text and Table S3.

The tachykinin system -controls
Raw Ct values were normalized to the internal reference gene and represented as delta Ct (dCT), exemplified for DLPFC and LC in the control male and female subjects (Fig. 2 A-I).Of note: Lower dCT values correspond to lower Ct values and thereby higher abundance of the gene of interest and vice-versa.The levels of all markers of the tachykinin system were in general higher in the DRN, LC and even MRN when compared to DLPFC and ACC, showing that these markers are more abundant in the lower brain stem than in cortical regions.Raw Ct values (rCT) for SP, TAC1 and their receptors (TACR1, TACR2 and TACR3) are shown in Table S4.The expression profile for TAC1 and SP were similar with the ranking order being DRN>LC>MRN>PFC=ACC.
Statistically significant sex differences were observed for TAC1 and TACR1 in the DLPFC with higher levels of TAC1 (p < 0.1) and TACR1 (p < 0.5) in the male controls (Fig. 2B and C).Fig. 2J shows a bubble plot for the dCT values of the tachykinin markers as a representation of the abundance of these genes in male and female controls.

Previously published studies.
Many studies describe the distribution of the substance P/tachykinin system in the human brain (e.g.(Ghatei et al., 1984;Bouras et al., 1986;Mai et al., 1986;Hornung et al.,  1992).Detailed studies have been published on the DRN and LC, whereby there are in the DRN different SP neuron populations, some expressing serotonin (Halliday et al., 1990;Baker et al., 1991;Fodor et al., 1992;Sergeyev et al., 1999).
The distribution of NK1-3 receptors have been studied in the human brain using various methodologies (Kus et al., 1998;Bensaid et al., 2001;Caberlotto et al., 2003;Rigby et al., 2005).Furthermore, the tachykinin system, especially the NK1 receptor, has been explored in particular in various forebrain areas of both men and women using positron emission tomography (PET) (Bergström et al., 2004;Hietala et al., 2005;Engman et al., 2012).Finally, many RNAseq and single-cell studies present information on the type of cortical neurons expressing tachykinin family transcripts (e.g.(Hodge et al., 2019).Our results are in agreement with their findings.
Taken together, our results support presence of moderate SP/TAC1 and TACR1-3 expression in different cortical regions with higher transcript levels in LC and even higher in DRN, of both sexes.
In the DRN only SP levels were significantly elevated, but only in male DSS (p < 0.05) (Table 2).In the LC, however, SP levels were significantly higher both in males and females (p < 0.05, Fig. 4A, F), versus TAC levels only in females (p < 0.05, Fig. 4G).Regarding receptors, TACR1 and TACR3 levels were significantly higher in both male and female DSS (p < 0.05, Fig. 4C, H and E, J), but TACR2 levels only in females (Fig. 4I, Table 2).

The neuropeptide Y system -controls
dCT values for the NPY system reveal a high abundance of NPY and NPYR1 in the DLPFC and ACC when compared to LC, DRN and MRN, associating these transcripts primarily with cortical systems.In contrast, NPYR2 and NPYR5 expression was lower with relatively similar dCT values in most of these regions (Table S5).
NPY levels were by far highest in the two cortical regions when compared to the brain stem regions and thus, the ranking order for NPY based on raw Ct values is DLPFC>ACC>>>LC>DRN>MRN (Fig. 2F-I, Table S5 for raw Ct values).
NPYR1 was most abundant in the DLPFC with the ranking order DLFPC>>ACC>LC>DRN>MRN, whereas NPYR2 levels were in general low: DLPFC=DRN>LC>ACC=MRN. NPYR5 was expressed at  approximately similar levels in all regions, except MRN (lower).
Regarding sex differences NPY transcript levels were significantly higher in female DLFPC (p < 0.5) and LC (p < 0.1), versus males (Fig. 2F, Table S5).Fig. 2J shows a bubble plot for the dCT values of the tachykinin and NPY markers as a representation of the abundance of these genes in male and female controls.
PET studies on the human NPY system are rare, focusing on the Y1 receptor (Hostetler et al., 2011), even if ligands have been generated (Fonseca et al., 2022).Also for the NPY system there are many RNAseq and single-cell studies with focus on cortex (e.g.(Hodge et al., 2019).

The neuropeptide Y systemdepressed subjects
In the DLPFC only the NPYR1 levels were significantly higher, both in male and female DSS (p < 0.05 for both), and in the LC, but only in male DSS (p < 0.05) (Fig. 5B, F, Table 2).NPY levels were significantly increased only in the LC, both of male and female DSS (p < 0.01 and p < 0.05 respectively, Fig. 5C, G, Table 2).

The CCK systemcontrols and depressed subjects
CCK levels were, like NPY, very high in cortex [PFC>ACC>>LC>>DRN>>MRN]. CCKBR levels were highest in cortex and LC with the ranking being PFC=ACC=LC>>DRN>>MRN.The levels of CCKAR were low, but highest in PFC and LC>>DRN>>MRN.There were no differences between male and females, nor between controls and DSS (Fig. S6).

LCM sequencing of isolated, human LC neurons
LCM-seq data showed presence of many neuropeptide families in noradrenergic LC neurons from 'normal' subjects including the tachykinin, NPY and CCK systems as well as the galanin system.Amino acid transmitter systems (VGLUT1-3 and GAD1-2) were also detected.

Discussion
Our study reports novel and marked regional differences in the mRNA expression of the TAC, NPY and CCK systems in five regions of human control and DSS postmortem brain tissue.In addition, significant alterations in expression of transcripts in DSS compared to controls were seen for the TAC and NPY but not for the CCK family (Table 2).For comparison, we include here (in the discussion) data on the galanin system from a previous study on the same samples (Barde et al., 2016) summarized in Table 2, allowing for three peptide systems to be overviewed (CCK is excluded from the table as no changes were recorded).
Since the study of DSS and controls was carried out on bulk samples, we do not know in which type of cells the transcripts are expressed.Therefore, the second part of our study (LCM-seq) on LC neurons isolated from 'normal' brains allows validation of at least this nucleus.Thus, we can show, using LCM-Seq, that 'normal' noradrenergic LC neurons express a large number of additional peptide and peptide receptor transcripts, in fact many at considerably higher levels than the four that we have focused on.Finally, we provide evidence that LC neurons may also perhaps also signal via glutamate and GABA, i.e. amino acid transmitters.

Normal distribution patterns and sex differences -bulk study
The present results confirm that the tachykinin system, in general, is strongly expressed in the lower brain stem, i.e.LC and especially in DRN, where TAC and SP as well as TACR1 and -R3 transcript levels are higher than in any other region.In contrast, both NPY and CCK are most highly expressed in the cortical regions, as are the NPYR1 and CCKBR transcript levels, although CCKB levels are high also in LC.Sex differences for controls are only found in two regions: TACR3 and NPY are higher in the female LC, and in DRN, NPYR2 is higher in female DRN.The GAL transcript (and galanin peptide), are most highly expressed in the LC sample (Barde et al., 2016).

The CCK, galanin, NPY and TAC systems
GAL and TACR1 were detected in all subjects using LCM-seq, whereas the other transcripts were only found on some.There was no obvious correlation observed between the expression pattern and age, sex or cause of death for the subjects.GalR3, NPY4R and CCKAR were not detected in any of the studied subjects.TAC1 (max ~11), NPY (~10.5) and GAL (~10) levels were high in some subjects.The large variation in counts for the ligands is in agreement with an in situ  , 2013).Possibly these differences indicate stages of activity: high transcript levels may reflect a compensatory increase in synthesis due to increased firing and neuropeptide release.The excitatory receptors (TACR1 and TACR3) are abundant, in all cells and fairly evenly expressed.Also, the expression of the inhibitory receptors (GALR1 and NPYR1) is robust, even if in fewer subjects.

Further peptide systems
The noradrenergic LC neurons express a large number of additional peptide and peptide receptor transcripts, sometimes at very high levels (and detected in most samples).It will be a major task to study the role of these neuropeptide systems in LC signalling, especially by applying electrophysiological methods as we and others have carried out with the galanin ligands.

Amino acid transmitter systems
Earlier animal experiments have shown that some catecholamine neurons in rodents express markers for glutamate, e.g.C1 adrenaline neurons (Guyenet et al., 2013) and mesencephalic A10 dopamine neurons (Descarries et al., 2008).We now report that some human LC neurons express VGLUTs and GAD.The results certainly suggest that LC neurons may not only signal via noradrenaline and neuropeptides but also amino acids, possibly of relevance for treatment of MDD with the glutamate antagonist ketamine (Kadriu et al., 2019;Krystal et al., 2019).

Type of neurons expressing transcripts and potential effects of ligands
Our bulk samples include several neuronal and non-neuronal cell population, and it is therefore important to define, in which population (s) the studied transcripts are expressed.Such information can be extracted from single-cell/nuclei studies e.g. for DLPFC (Mathys et al., 2019;Nagy et al., 2020) and DRN and LC (Weber et al., 2022) and are provided for LC in the present LCM-seq study data.In the following paragraphs (4.3.1., 4.3.3. and 4.3.5.) we will rely on data from these studies.Regarding potential functional effects of peptides we have mainly to refer to results from animal experiments.

Cortical phenotypes
TAC1, TAC3 and NPY are primarily localized in GABAergic neurons, but NPY also in glutamatergic neurons and CCK in both types of neurons.TACR1 is mainly expressed in GABAergic neurons; TACR3 mainly in glutamatergic neurons; NPY1R in GABAergic and glutamatergic neurons and NPY5R in glutamatergic neurons; CCKB in glutamatergic neurons.

DRN phenotypes
The 5-HT neurons are characterized by (i) a robust expression of TAC1 and a modest signal for TACR1 and TACR3; (ii) in general a very low expression of the NPY family members -only NPYR1 is detected; and (iii) a weak expression of GalR2 and GALR3, but no signal for GAL or GALR1.All transcripts are in addition expressed at various levels in excitatory and inhibitory neurons, as well as in other cell types.

DRN functions
In rat, intracellular recordings from 5-HT neurons show a robust increase in excitatory postsynaptic currents after administration of either SP (TACR1 agonist) or senktide (NKB; TACR3 agonist); however, this effect is not direct but is mediated via stimulation of, possibly local, glutamatergic neurons (Liu et al., 2002).Galanin has been reported to hyperpolarize most 5-HT-neurons in the most dorsal part of the nucleus (Xu et al., 1998).However, it has also been reported that galanin in the DRN acts via GalR1 on GABA interneurons and via GalR2 on 5-HT neurons (Sharkey et al., 2008).

LC phenotypes
Our own analysis on isolated LC neurons shows that TAC1, NPY, CCK and GAL are robustly expressed (GAL>TAC1=NPY) in noradrenergic LC cells, as are TACR1 and TACR3, and to a lesser extent CCKBR and GALR1 (but not CCKAR).NPYR transcripts are only detected in a few cells or not at all.These results agree with the above mentioned single-nucleus paper (Weber et al., 2022).

LC functions
In rodents, SP has a direct excitatory effect on noradrenergic LC neurons (Guyenet and Aghajanian, 1977;Engberg et al., 1981).In contrast, NPY has a depressant effect (Olpe and Steinmann, 1991) and potentiates the NA-induced inhibition, via NPYR2 (Illes et al., 1993).Several studies have shown that galanin reduces firing of rat LC neurons (see (Pieribone et al., 1995)) and mouse neurons (Caramia et al., 2023).Also, how the co-existing amino acids glutamate and GABA transmitters participate in signalling of noradrenergic LC neurons remains to be analysed.

Tachykinin and NPY (and galanin) systems are changed in MDD
The present and a previous study suggest, as indicated in Table 2, that three neuropeptide systems, SP/TAC, NPY and galanin, are associated with MDD, and mainly in two regions: DLPFC and LC.No differences between controls and DSS were observed in the CCK system (Table S6).This may not be surprising, since this system has mainly been associated with panic disorder and anxiety (Rehfeld, 2000;Harro, 2006;Zwanzger et al., 2012); however, in the frontal cortex, a significantly higher number of CCK receptors and affinity constants were found in a series of suicide victims (Harro et al., 1992).
Regarding DLFPC in DSS both ligands and receptors of the SP/TAC and galanin systems are upregulated (Table S4), mainly in females, possibly in agreement with the notion that women are more likely to be afflicted by MDD (Kessler et al., 2003).The binding of [125I]BH-SP in orbitofrontal cortex (BA 47) is decreased in MDD (Stockmeier et al., 2002), apparently contrasting our data.Regarding the NPY system, increased NPYR2 levels have been reported in layer IV in DSS (Caberlotto and Hurd, 2001), as well as upregulation of NPYR1, NPYR2 and decreased NPY transcript levels in female DSS (Sharma et al., 2022), whereas we only detected upregulation of NPYR1.However, GAL and GALR3 are downregulated in males.
Regarding the galanin system in the human PFC, RNAScope analysis from our previous study shows that GAL, GALR1 and -R3 transcripts are mainly expressed in glutamatergic projection neurons (Zhong et al., 2022).We have hypothesized that galanin via the inhibitory GalR3 on intra-telencephalic pyramidal neurons protects against glutamatergic over-excitation, whereas GALR1 in pyramidal neurons may instead influence thalamic signalling.Thus, the galanin system may offer a further target for ketamine MDD therapy (Kaudri et al., 2019;Krystal et al., 2019).Overall galanin may possibly be of significance for neuroprotection and resilience (see Figs. 6 and S6 in (Zhong et al., 2022).SP, TAC and NPY are, on the other hand and as said, mainly expressed in GABAergic interneurons (DeFelipe et al., 2013).
Our results indicate that the LC is the most strongly affected region in MDD, with increased levels of several transcripts of the three peptide systems, in both sexes.The increase of SP transcript may reflect increased SP release that, together with increased availability of TACR1 receptors, may lead to activation of the noradrenergic neurons.This likely elevated excitation by the tachykinin system may be counteracted by the primarily inhibitory peptides such as NPY and galanin.In fact, both galanin (Sciolino et al., 2015;Hökfelt et al., 2018) and especially NPY (Heilig, 2004;Morales-Medina et al., 2010;Cohen et al., 2012;Sabban et al., 2016;Kautz et al., 2017) have at the level of the LC been suggested to be involved in resilience.We hypothesize that in MDD the excitatory, prodepressive drivers like tachykinins, corticotropin releasing hormone (CRH) and hypocretin/orexin have 'won over' anti-depressive, 'resilience-inducing' factors like NPY and galanin.

Limitations
The present study only reports on transcripts, i.e. it lacks information on peptides/proteins, However, we cite numerous reports based on radioimmunoassay or immunohistochemistry, evidencing presence of the translated proteins.The long storage of our present samples compared to the study by (Barde et al., 2016) does not seem to matter since, e.g.NPY mRNA levels (still) are very high in cortex.Please, note that Barde et al. (2016) is a more complex study, encompassing both radioimmunoassay and DNA methylation.In the present Study #1 on bulk samples we do not know about the phenotype of the cells expressing peptide systems.However, thanks to the LCM-seq analysis (i. e. Study#2) we know that at least the four peptide systems (CCK, galanin, NPY and TAC/substance P) are expressed in noradrenergic LC neurons.Two control subjects show presence of SSRIs at the time of death.SSRIs are now often prescribed for many conditions that do not fall into Axis 1 categories.Therefore, two subjects were not depressed but did take SSRIs and hence placed in the control group.It is very difficult to completely avoid such factors in postmortem studies, and a future aim is to collect more samples with such treatments (to increase 'n' and statistical power) and check, if there is any possible influence on expression of these genes.The LCM-seq study was carried out on post-mortem tissue from 20 subjects of different ages and is based on pooling of many neurons (~50-120 neurons) from the same subject.Although this should increase sensitivity, conclusions on transcript levels of individual neurons or on a possible co-existence of transcripts cannot be drawn.Finally, we have in several cases speculated about the role of the peptide systems in MDD: further ISH analysis and immunohistochemistry (when receptor antibodies become available) would allow validation of such hypotheses.

Fig. 2 .
Fig. 2. Expression of the tachykinin and NPY systems in the dorsolateral prefrontal cortex (DLPFC) and locus coeruleus (LC) of male and female controls.Raw CT values for the markers were normalised to an internal reference gene and shown as dCT values for the two regions (A-I) and a bubble plot shows the overview of the delta CT values (J).Sex differences in the expression levels were observed for TAC1 and TACR1 in the DLPFC and for TACR3 and NPY in the LC.*P <0.05, **p < 0.01.Abbreviations: CT: cycle threshold, dCT: delta CT, NPY: Neuropeptide Y, NPYR1: Neuropeptide receptor 1, NPYR2: Neuropeptide receptor 2, NPYR5: Neuropeptide receptor 5, TAC1: Tachykinin 1, TACR1: Tachykinin receptor 1, TACR2: Tachykinin receptor 2, TACR3: Tachykinin receptor 3.

Fig. 6 .
Fig. 6.LCM-seq data for galanin, NPY and TAC family and their receptors as well as VGLUT2, GAD2 and CARTPT from isolated neurons of the human LC.RNA sequencing data analysed by log2 transformation of normalized data.Normalized counts for TAC and its receptors (A), NPY and its receptors (B), CCK family (C), galanin and its three receptors (D), VGLUT2 and GAD (E) and CARTPT (F) is shown.List of peptides (H) and peptide receptor (I) transcripts with cut off value of ≥1.0, ≥5.0 and ≥10.0.